A3.1-1

Appendix 3.1

Evidence Tables

References A3.1-80

List of Tables

Table A3.1-1 Studies on aerosolized nicotine and dependence, by dependency criteria A3.1-2

A. A strong desire or sense of compulsion to take the substance A3.1-2

B. A persistent desire or unsuccessful efforts to reduce or control substance use A3.1-2

C. A physiological withdrawal state when substance use is reduced or ceased A3.1-3

D. Use the same (or closely related) substance with the intention of relieving or avoiding withdrawal symptoms

A3.1-3

E. Tolerance to the effects of the substance A3.1-6

F. Preoccupation with substance use, as manifested by important alternative pleasures or interests being given up

or reduced because of substance use, or by spending a great deal of time in activities necessary to obtain and

take the substance or recover from its effects A3.1-7

G. Persistent substance use despite clear evidence of harmful consequences A3.1-7

Table A3.1-2 Human studies on the effects of nicotine exposure on adolescent users A3.1-8

Table A3.1-3 Preclinical/animal studies on adolescent nicotine exposure A3.1-18

Table A3.1-4 Human studies on the effects of nicotine exposure on fetal brain development A3.1-33

A. Sudden infant death syndrome and potentially related risk factors A3.1-33

B. Altered corpus callosum A3.1-34

C. Auditory processing defects A3.1-35

D. Appetitive behaviors: Smoking uptake, nicotine dependence A3.1-40

E. Appetitive behaviors: Use of other substances A3.1-48

F. Appetitive behaviors: Overweight and obesity A3.1-51

G. Appetitive behaviors: Altered size and sensitivity of brain reward systems A3.1-54

H. Attention and cognition, behavioral disorders A3.1-56

Table A3.1-5 Preclinical/animal studies on fetal nicotine exposure A3.1-63
A Report of the Surgeon General

A3.1-2 Appendix 3.1

Table A3.1-1 Studies on aerosolized nicotine and dependence, by dependency criteria

A. A strong desire or sense of compulsion to take the substancea

Study Design/population Findings

McQueen et al. (2011) • Interviews with people at MidWest Vapefest

• Meetings with members of MidWest Vapers 

Group

• Sample size = 15

• Participants reported not “having the same sense of 

urgency about vaping that I had about smoking. . . . I go 

all day without vaping, and it doesn’t occur to me.”

Dawkins et al. (2013) • Survey of users of two popular brands of 

e-cigarettes in the United Kingdom

• Sample size = 1,347

• 18% of current and former smokers craved e-cigarettes 

as much as tobacco.

• 23% used an e-cigarette within 5 minutes of waking.

• 49% used e-cigarettes within 6–30 minutes of waking.

Goniewicz et al. (2013) • Survey of e-cigarette users in Poland

• Sample size = 179

• 98% used e-cigarettes every day.

• 44% used e-cigarettes within 30 minutes of waking.

Etter (2015) • Cross-sectional survey in English and 

French of daily e-cigarette users who had 

quit smoking in the previous 2 months

• Sample size = 374

• 38% reported “definitely” and 23% “a lot” about having 

strong urges to use e-cigarettes today.

• 68% reported “definitely” feeling an irresistible urge 

to use e-cigarettes “after a few hours without using an 

e-cigarette.”

• Perceived effect of e-cigarettes on craving to smoke was 

associated with dependence on e-cigarettes and time to 

first puff of e-cigarettes after waking.

Etter and Eissenberg 

(2015)

• Survey of daily e-cigarette users from 

outside the United States

• Sample size = 1,284

• 30.7% of people who used e-cigarettes containing 

nicotine reported they likely would not be able to stop 

using e-cigarettes. 

• 28.2% reported that it would be “very difficult” or 

“impossible” to stop using e-cigarettes. 

• 27.5% reported not being able to stop e-cigarette use.

Foulds et al. (2015) • Survey of e-cigarette users who were 

former cigarette smokers

• Sample size = 3,609

• 35% had strong cravings to use e-cigarettes.

• 6.8% woke up at night to use e-cigarettes.

• 93% had strong cravings to smoke a conventional 

cigarette.

• 41.2% reported waking up at night to smoke cigarettes 

prior to quitting.

aMost of the evidence for this criterion comes from subjective reports of users regarding their experiences with e-cigarettes.

B. A persistent desire or unsuccessful efforts to reduce or control substance useb

Study Design/population Findings

Dawkins et al. 

(2013)

• Survey of users of two popular brands 

of e-cigarettes in the United Kingdom

• Sample size = 1,347

• 12% of people who were attempting to reduce e-cigarette use 

reported being “very” or “extremely” successful.

Etter and 

Eissenberg (2015)

• Survey of daily e-cigarette users from 

outside the United States

• Sample size = 1,284

• Of people who used e-cigarettes containing nicotine: 

– 30.7% reported they would likely not be able to stop using 

e-cigarettes. 

– 28.2% reported that it would be “very difficult” or “impossible” 

to stop using e-cigarettes. 

– 27.5% reported not being able to stop e-cigarette use.

bAs applied to the use of e-cigarettes, there is little support for this criterion because of limited data on users of e-cigarettes trying 

to quit e-cigarette use. Many people report using e-cigarettes to replace, reduce, or avoid relapsing to cigarette use (Etter and Bullen 

2011; Adkison et al. 2013; Dawkins et al. 2013; Vickerman et al. 2013; Etter and Eissenberg 2015) and may have no intention of stopping all nicotine or tobacco use.
Evidence Tables A3.1-3

E-Cigarette Use Among Youth and Young Adults

C. A physiological withdrawal state when substance use is reduced or ceasedc

Study Design/population Findings

Vansickel and Eissenberg 

(2013)

• Experienced users of e-cigarettes 

who puffed in one session with 

four phases: baseline, 10-puff bout, 

ad libitum bout, and rest period

• Sample size = 8

• Significant reductions in anxiety, “restlessness,” 

and intention to smoke from abstinence to 

post-electronic-cigarette use.

• Significant increases in “feel awake,” “calm you down,” and 

“concentrate” from abstinence to post-electronic-cigarette 

use.

Dawkins and Corcoran 

(2014)

• Regular users of e-cigarettes who 

engaged in 10- and 60-minute 

puffing bouts with e-cigarettes

• Sample size = 14 

• Significantly higher ratings of “urge to smoke” and nicotinerelated withdrawal symptoms at tobacco-abstinent baseline 

compared with post-electronic-cigarette use.

Spindle et al. (2015) • Experienced users of e-cigarettes 

who preferred two sessions with 

10-puff bouts

• Sample size = 13

• Significant reductions in “anxious,” “urge to use an 

e-cigarette,” “craving an e-cigarette,” and “impatient” from 

tobacco-abstinent baseline to post-electronic- cigarette use.

• Increases in “awake,” “calm,” and “concentrate” from 

tobacco-abstinent baseline to post-electronic-cigarette use.

St. Helen et al. (2016) • E-cigarette users recruited over 

the Internet participated in a 1-day 

research ward study. Subjects took 

15 puffs from their usual brand 

of e-cigarette. Exhaled breath was 

trapped in gas-washing bottles, 

and blood was sampled before and 

several times after use.

• Thirteen healthy, experienced adult 

e-cigarette users (6 females and 

7 males).

E-cigarettes delivered a mean of 1.3 mg (95% CI, 0.9–1.8) 

of nicotine, and 94% of the inhaled dose, 1.2 mg (0.8–1.7), 

was systemically retained. Mean maximum plasma nicotine 

concentration (Cmax) was 8.4 ng/mL (95% CI, 5.4–11.5) and 

time of maximal concentration (Tmax) was 2 to 5 minutes; one 

participant had Tmax of 30 minutes. In this study, 89% and 

92% of vegetable glycerin and propylene glycol, respectively, 

was systemically retained. Heart rate increased by an average 

of 8.0 beats per minute after 5 minutes. Withdrawal and urge 

to smoke decreased, and the e-cigarettes were described as 

satisfying.

cResults of these studies may be confounded by the use of conventional cigarettes by many users of e-cigarettes.

D. Use the same (or closely related) substance with the intention of relieving or avoiding withdrawal symptoms

Study Design/population Findings

Bullen et al. (2010) • Single-blind, randomized, repeated-measures, 

crossover trial of users of the Ruyan V8 

e-cigarette in New Zealand

• Dependent smokers were randomized to 

16 mg or 0 mg nicotine e-cigarettes.

• Sample size = 40

• Significant reduction in desire to smoke for users of 

16 mg nicotine e-cigarettes compared with users of 

0 mg nicotine e-cigarettes. 

• Other withdrawal symptoms—such as irritability, 

restlessness, and poor concentration—were not 

dependent on dose.

Vansickel et al. (2010) • Smokers who never used e-cigarettes who 

engaged in two 10-puff bouts of four Latinsquare order conditions: own-brand cigarette, 

NPRO e-cigarette, Hydro e-cigarette, and sham 

cigarette

• Sample size = 32

• Two brands of e-cigarettes significantly decreased 

ratings of tobacco abstinence symptoms, but to a 

lesser magnitude than conventional cigarettes.

Table A3.1-1 Continued
A Report of the Surgeon General

A3.1-4 Appendix 3.1

Study Design/population Findings

Etter and Bullen 

(2011)

• Survey in English and French of visitors 

to websites and Internet forums related to 

e-cigarettes and smoking cessation

• Sample size = 3,587

• 50% reported that e-cigarette use may relieve 

cravings where smoking is prohibited.

• 60% reported that e-cigarettes may satisfy the desire 

to smoke.

• 51% reported that e-cigarette use may help with 

quitting smoking.

• 55% reported that e-cigarette use may help with 

reducing cigarette smoking.

• 79% reported using e-cigarettes to deal with cravings 

for tobacco.

• 67% reported using e-cigarettes to deal with 

withdrawal symptoms.

• 77% reported using e-cigarettes to help with 

quitting smoking or avoiding relapse to conventional 

cigarettes.

Adkison et al. (2013) • Data from Wave 8 of the International Tobacco 

Control Four-Country Survey

• Sample size (had tried an e-cigarette) = 450 

• 75.4% reported using e-cigarettes to help reduce 

smoking.

• 85.1% reported using e-cigarettes to help quit 

smoking.

Bullen et al. (2013) • Randomized, controlled, superiority trial in 

Auckland, New Zealand

• Sample size = 657 

• People were randomized (289 to nicotine 

e-cigarettes, 295 to patches, and 73 to 

placebo e-cigarettes) and were included in the 

intention-to-treat analysis.

• At 6 months, verified abstinence was

– 7.3% (21 of 289) with nicotine e-cigarettes;

– 5.8% (17 of 295) with patches; and

– 4.1% (3 of 73) with placebo e-cigarettes. 

• Risk difference for nicotine e-cigarette vs. patches 

1.51 [95% CI, −2.49 to 5.51]; for nicotine e-cigarettes 

vs. placebo e-cigarettes 3.16 [95% CI, −2.29 to 8.61]).

• No significant differences in adverse events between 

products.

Caponnetto et al. 

(2013) 

• 12-month randomized controlled trial of 

smoking reduction/abstinence in smokers not 

in the process of quitting

• Sample size = 300 

• Group 1 received 7.2 mg nicotine e-cigarettes 

for 12 weeks.

• Group 2 received 7.2 mg nicotine e-cigarettes 

for 6 weeks followed by 5.4 mg nicotine 

e-cigarettes for 6 weeks.

• Group 3 received placebo (no nicotine) 

e-cigarettes for 12 weeks.

• Declines in cig/day use and levels of exhaled carbon 

monoxide were observed at each study visit in all 

three study groups.

• In smokers not intending to quit, the use of 

e-cigarettes, with or without nicotine, decreased 

cigarette consumption and elicited enduring tobacco 

abstinence without causing significant side effects.

Dawkins et al. (2013) • Survey of users of two popular brands of 

e-cigarettes in the United Kingdom

• Sample size = 1,347

• 76% reported initiating e-cigarette use as a “complete 

alternative to smoking.”

• 67% reported wanting a “complete alternative to 

smoking.”

• 7% reported initiating e-cigarette use to quit 

smoking.

• E-cigarette use was associated with decreased 

cravings for tobacco cigarettes: 91% reported 

substantial decreases in cravings for conventional 

cigarettes.

Table A3.1-1 D Continued
Evidence Tables A3.1-5

E-Cigarette Use Among Youth and Young Adults

Study Design/population Findings

Dockrell et al. (2013) • Survey of smokers in Great Britain who had 

and had not used e-cigarettes

• Sample size = 1,380

• 50% reported that e-cigarette use may relieve 

cravings where smoking is prohibited.

• 60% reported that e-cigarette use may satisfy the 

desire to smoke. 

• 51% reported that e-cigarette use may help with 

quitting smoking.

• 55% reported that e-cigarette use may help with 

reducing cigarette smoking.

Vansickel and 

Eissenberg (2013)

• Experienced users of e-cigarettes who puffed 

in one session with four phases: baseline, 

10-puff bout, ad libitum bout, and rest period

• Sample size = 8

• Significant reductions in “anxious,” “restlessness,” 

and intention to smoke from abstinence to postelectronic-cigarette use.

• Significant increases in “feel awake,” “calm you 

down,” and “concentrate” from abstinence to postelectronic-cigarette use.

Vickerman et al. 

(2013)

• Survey of quitline users in six states

• Sample size (ever users of e-cigarettes) = 765

• 51% reported using e-cigarettes to help quit other 

types of tobacco use.

• 15% reported using e-cigarettes to replace other 

types of tobacco use.

• 7% reported using e-cigarettes to reduce other types 

of tobacco use.

Dawkins and Corcoran 

(2014)

• Regular users of e-cigarettes who engaged 

in 10- and 60-minute puffing bouts with 

e-cigarettes

• Sample size = 14

• Significant reductions in urge to smoke and nicotinerelated withdrawal symptoms after 10- and 60-minute 

puff periods.

Grana et al. (2014) • Survey of current smoking in the United 

States

• Sample size = 949

• E-cigarette use at baseline did not significantly 

predict quitting 1 year later.

• E-cigarette use by smokers was not followed by 

greater quitting.

Norton et al. (2014) • Smokers who abstained from conventional 

cigarettes and used only e-cigarettes for 

72 hours

• Lab-monitored e-cigarette puffing sessions 

that occurred before and after the 72-hour 

period

• Sample size = 32

• Initial use of e-cigarettes did not significantly reduce 

scores on either scale of the Questionnaire on 

Smoking Urges.

Wagener et al. (2014) • E-cigarette-naïve smokers sampled three 

e-cigarettes for a <10-minute puffing bout, 

followed by ad libitum e-cigarette use for 

1 week.

• Sample size = 19

• No significant changes in nicotine withdrawal 

symptoms from baseline to post-electronic-cigarette 

use.

Etter (2015) • Cross-sectional survey in English and French 

of daily e-cigarette users who had quit 

smoking in the previous 2 months

• Sample size = 374

• 25% reported that addiction to e-cigarettes is the 

same strength as addiction to conventional cigarettes.

• 2% reported that addiction to e-cigarettes is stronger 

than addiction to conventional cigarettes.

Etter and Eissenberg 

(2015)

• Survey of daily e-cigarette users from outside 

the United States

• Sample size = 1,284

• 65% of users of e-cigarettes with nicotine reported 

that it is “very true” or “extremely true” that they use 

e-cigarettes to deal with cravings for cigarettes.

• 82% reported using e-cigarettes to help with 

quitting smoking or avoiding relapse to conventional 

cigarettes.

Table A3.1-1 D Continued
A Report of the Surgeon General

A3.1-6 Appendix 3.1

Study Design/population Findings

Lechner et al. (2015) • Current smokers were randomized to a 

crossover design using a first- and secondgeneration e-cigarette on two separate days.

• Sample size = 22

• Significant reductions in nicotine withdrawal 

symptoms after e-cigarette use.

• Greater reductions in nicotine withdrawal symptoms 

after use of the 2nd-generation e-cigarette compared 

with the 1st-generation e-cigarette.

Spindle et al. (2015) • Experienced users of e-cigarettes who 

preferred two sessions with 10-puff bouts

• Sample size = 13

• Significant reductions in “anxious,” “urge to use an 

e-cigarette,” “craving an e-cigarette,” and “impatient” 

from tobacco-abstinent baseline to post-electroniccigarette use.

• Increases in “awake,” “calm,” and “concentrate” 

from tobacco-abstinent baseline to post-electroniccigarette use.

E. Tolerance to the effects of the substance

Study Design/population Findings

Farsalinos et al. (2013) • Interviews with experienced users of 

e-cigarettes who had completely substituted 

e-cigarettes for conventional cigarettes for at 

least 1 month

• Sample size = 111

• 16.2% reported increasing the level of nicotine in 

e-cigarette liquid to achieve smoking abstinence.

• No users reported decreasing the level of nicotine in 

e-cigarette liquid before smoking cessation.

• The majority of e-cigarette users also used 

conventional cigarettes (Sutfin et al. 2013; Aboaziza 

and Eissenberg 2015; Barnett et al. 2015) and were 

likely already dependent on nicotine.

Polosa et al. (2014) • Prospective observational study in Italy, 

evaluating smoking reduction or abstinence 

in smokers not intending to quit using an 

e-cigarette

• 6-month intervention using e-cigarettes, with 

follow-ups through 24 months

• Sample size = 24

• Four participants upgraded their initial e-cigarette to 

a higher performing e-cigarette.

Table A3.1-1 D Continued
Evidence Tables A3.1-7

E-Cigarette Use Among Youth and Young Adults

F. Preoccupation with substance use, as manifested by important alternative pleasures or interests being given up or 

reduced because of substance use, or by spending a great deal of time in activities necessary to obtain and take the 

substance or recover from its effectsd

Study Design/population Findings

Etter and Bullen (2011) • Survey in English and French of visitors 

to websites and Internet forums related to 

e-cigarettes and smoking cessation

• Sample size = 3,587

• 39% reported using e-cigarettes to deal with 

situations in which smoking is not allowed.

Dawkins et al. (2013) • Survey of users of two popular brands of 

e-cigarettes in the United Kingdom

• Sample size = 1,347

• 13% reported using e-cigarettes more than 

conventional cigarettes because e-cigarette use is 

permitted in places where smoking is not.

Vickerman et al. (2013) • Survey of quitline users in six states

• Sample size (ever users of e-cigarettes) = 765

• 5.4% reported using e-cigarettes in places where 

tobacco is not allowed.

Mimms (2014) • Article from onvaping.com posted in Vaping 

Tips

• Text and videos explain how to use the “stealth 

vaping” technique.

Bohrer (2015) • Article from vapenewsmagazine.com • “Stealth vaping” is a style of e-cigarette use that is 

not easily detected by others.

dCriterion may be slightly less relevant for nicotine than other substances of abuse.

G. Persistent substance use despite clear evidence of harmful consequencese

Study Design/population Findings

Adkison et al. (2013) • Data from Wave 8 of the International 

Tobacco Control Four-Country Survey

• Sample size (had tried an e-cigarette) = 450

• 66–82% of respondents who were aware of 

e-cigarettes believed that e-cigarettes are less 

harmful than conventional cigarettes.

Dockrell et al. (2013) • Survey of smokers in Great Britain who had 

and had not used e-cigarettes

• Sample size = 1,380

• 71% of smokers believed that e-cigarettes are safer 

than tobacco cigarettes.

• 28% of smokers believed that e-cigarettes are safer 

than nicotine replacement therapies.

Goniewicz et al. (2013) • Survey of e-cigarette users in Poland

• Sample size = 179

• 82% did not think that e-cigarettes are completely 

safe but thought that they were less dangerous than 

conventional cigarettes.

Carroll Chapman and 

Wu (2014)

• Review of articles focused on e-cigarette use

• Sample size = 21 studies

• 54% believed e-cigarettes are safer than tobacco.

Note: CI = confidence interval; mg = milligram.

eAs applied to the use of e-cigarettes, there is little support for this criterion because of limited data on the health effects of e-cigarette 

use. Notably, e-cigarette use is often associated with replacing or reducing the use of conventional cigarettes (Etter and Bullen 2011; 

Adkison et al. 2013; Dawkins et al. 2013; Vickerman et al. 2013; Etter and Eissenberg 2015). Although various potentially harmful 

ingredients have been identified in e-cigarettes, their levels may be substantially lower than those found in conventional cigarettes 

(McAuley et al. 2012; Goniewicz et al. 2014a,b; Tayyarah and Long 2014). Additionally, some people who have replaced at least some 

cigarettes with e-cigarettes report improvements in their health (McQueen et al. 2011; Dawkins et al. 2013; Farsalinos et al. 2013; 

Hua et al. 2013; Polosa et al. 2014; Etter 2015). 

Table A3.1-1 Continued
A Report of the Surgeon General

A3.1-8 Appendix 3.1

Table A3.1-2 Human studies on the effects of nicotine exposure on adolescent users

Study Design/population Outcomes examined Findings

Kandel et al. 

(1992)

• Representative cohort 

study

• 1,160 young adults in 

grades 10 and 11 in 

public high schools in 

the state of New York 

• Followed at intervals and 

ending 18–19 years after 

baseline (to 35 years of 

age)

Personal interviews were conducted 

to assess licit and illicit drug use and 

psychotropic drug use. The sequence of 

progression was analyzed.

Four stages of progression were 

identified: legal drugs (alcohol or 

cigarettes); marijuana; illicit drugs 

other than marijuana; and medically 

prescribed psychotropic drugs. 

Progression to illicit drug use among 

males was dependent on prior alcohol 

use. Progression to marijuana use 

among females was dependent on either 

cigarette or alcohol use. Age of onset 

and frequency of use at lower stages 

of drug use were strong predictors of 

progression.

Lewinsohn et al. 

(1999) 

• Longitudinal cohort 

study

• 684 high school students, 

14–18 years of age, from 

nine high schools in 

western Oregon

• Followed to age 24

Participants were randomly selected 

and evaluated for alcohol, cannabis, and 

other drug use or dependence. Lifetime 

tobacco use, age at onset of smoking, 

and frequency or quantity of smoking 

and quit efforts during adolescence were 

assessed through interviews. Diagnosis 

of substance abuse or dependence was 

made using DSM-IV.

Lifetime smoking among older 

adolescents was significantly associated 

with future alcohol, cannabis, illicit 

drug, and multiple drug use disorders 

during young adulthood. Being a former 

smoker did not reduce the risk of 

future substance use disorders but was 

associated with a reduced risk of alcohol 

use disorder. Among daily smokers, 

earlier age at onset of smoking predicted 

future substance use disorders. 

DiFranza et al. 

(2000)

• Longitudinal cohort 

study

• 681 students, 12–13 

years of age, in 7th grade 

in seven public 

schools in two cities in 

Massachusetts

• Followed for 1 year

Interviews were conducted to collect 

detailed information about tobacco use. 

The latency time to onset of symptoms 

of nicotine dependence was measured 

from the time a student first smoked at 

least once per month. 

Ninety-five students initiated occasional 

smoking; 22% of them (n = 21) reported 

symptoms of nicotine dependence 

within 4 weeks of initiating occasional 

smoking. Sixty (63%) reported one or 

more symptoms of nicotine dependence; 

62% of them (37) reported symptoms of 

dependence before smoking daily. 

Lai et al. (2000) • Population-based survey

• 17,809 respondents, 

12 years of age or 

older, from the 

NHSDA (U.S. civilian, 

noninstitutionalized 

population) 

Univariate and multiple logistic 

regression analyses were used to assess 

independent associations between 

cigarette smoking and use of other 

drugs. Results were adjusted for race 

and gender and stratified by age. The 

study excluded those with illicit drug 

use before smoking. 

Participants who smoked cigarettes were 

significantly more likely than never 

smokers to use cocaine, heroin, crack, 

and marijuana, even after adjusting for 

potential confounders. 
Evidence Tables A3.1-9

E-Cigarette Use Among Youth and Young Adults

Study Design/population Outcomes examined Findings

Hanna et al. 

(2001)

• Participants 12–16 years 

of age in the NHANES 

III (U.S. civilian, 

noninstitutionalized 

population)

Associations between smoking 

and other drug or alcohol use and 

behavior problems were assessed 

using multivariate logistic regression, 

controlling for potential confounders. 

Smoking status, age of onset of regular 

smoking, alcohol use, and illicit drug 

use were determined from interviews. 

Outcomes assessed included depression 

or dysthymia, on the basis of DSM-III 

criteria; school problems, including 

repeating grades, suspension, or 

expulsion; use of birth control pills or 

pregnancy; and psychosocial skills. 

Onset of regular smoking at 13 years 

of age or younger was associated with 

difficulty getting along with peers. 

Regular smoking was associated with 

alcohol use, illicit drug use, and school 

problems, regardless of onset of regular 

smoking. Smoking was not associated 

with depression or dysthymia but was 

associated with pregnancy. 

DiFranza et al. 

(2002)

• Longitudinal cohort 

study

• 679 urban 7th grade 

students, 12–13 years of 

age, from seven schools 

in two cities in central 

Massachusetts

• Followed for 1 year

Interviews were conducted to collect 

detailed information about tobacco use. 

The latency time to onset of symptoms 

of nicotine dependence was measured 

from the time a student first smoked at 

least once per month.

Of 332 students who used tobacco, 

40% reported dependence symptoms. 

The median latency from onset of 

monthly smoking to dependence 

symptoms was 21 days for girls and 

183 days for boys. Median frequency of 

use at the onset of symptoms was two 

cigarettes, 1 day per week. The presence 

of dependence symptoms predicted 

continued smoking. No minimum dose 

or duration of nicotine use required for 

symptoms was identified. 

Wagner and 

Anthony (2002)

• Population-based survey

• 44,624 U.S. 

household civilian, 

noninstitutionalized 

residents 12–25 years 

of age

Two mechanisms were studied to help 

account for prior observations about the 

“stepping-stone” or “gateway” sequences 

that link the use of alcohol, tobacco, 

marijuana, and cocaine. 

Users of tobacco and alcohol were 

more likely than nonusers to have an 

opportunity to try marijuana and more 

likely to use marijuana if given the 

opportunity. 

Isensee et al. 

(2003)

• Longitudinal study

• Analysis included 

3,021 randomly 

selected subjects, 

14–24 years of age at 

baseline, from the Early 

Developmental Stages of 

Psychopathology Study 

from Munich, Germany. 

• Follow-up = 4 years

Baseline and 4-year follow-up data were 

used to assess bidirectional associations 

between smoking and nicotine 

dependence and panic and other anxiety 

disorders. Cox regression with timedependent covariates was used to adjust 

for potential confounders. Diagnostic 

assessments were on the basis of the 

Munich-Composite International 

Diagnostic Interview. Panic attack and 

panic disorder, social phobia, phobia not 

otherwise specified, posttraumatic stress 

disorder, generalized anxiety disorder, 

agoraphobia, and obsessive-compulsive 

disorder were defined according to 

DSM-IV criteria. 

Panic attacks and panic disorder were 

strongly associated with occasional 

and regular smoking and nicotine 

dependence. Prospective analyses found 

significant associations between regular 

smoking and nicotine dependence and 

new-onset panic attacks and between 

nicotine dependence and onset of panic 

disorder. In regression analysis with 

adjustment for confounders, nicotine 

dependence was associated with panic 

attacks but not panic disorder. 

Table A3.1-2 Continued
A Report of the Surgeon General

A3.1-10 Appendix 3.1

Study Design/population Outcomes examined Findings

O’Loughlin et 

al. (2003)

• Longitudinal cohort 

study

• 1,267 students in 

7th grade who were 

enrolled in the McGill 

University Study on 

the Natural History of 

Nicotine Dependence

• Convenience sample 

from seven English and 

three French secondary 

schools in Montreal, 

Canada

• Followed for 6 years

Associations between five indicators 

of nicotine dependence and monthly, 

weekly, and daily smoking vs. less 

frequent smoking were assessed. Of the 

five indicators of nicotine dependence, 

one was based on the criteria for tobacco 

dependence in the ICD-10 and one on 

the HONC, and the remainder included 

three clusters of symptoms. 

More than 16% of past 3-month 

smokers, more than 19% of weekly 

smokers, and nearly 70% of daily 

smokers were tobacco-dependent. 

Nicotine dependence and cravings were 

significantly associated with frequency 

of smoking. 

AudrainMcGovern et al. 

(2004)

• Longitudinal cohort 

study

• 1,123 students in 

9th grade in public high 

schools in northern 

Virginia

• Followed through the 

end of 12th grade

Data were collected on smoking 

practices, novelty seeking, academic 

performance, substance use, peer 

smoking, physical activity, sports 

participation, and receptivity to tobacco 

advertisements.

Four smoking trajectories 

were identified: never smokers, 

experimenters, earlier/faster smoking 

adopters, and later/slower smoking 

adopters. Earlier/faster smoking 

adopters were characterized by 

high novelty-seeking personality, 

depressive symptoms, poorer academic 

performance, and receptivity to tobacco 

advertising.

John et al. 

(2004)

• Population-based study

• 1,601 current and 

836 former smokers, 

18–64 years of age, from 

northern Germany

Among adult smokers, this study 

assessed the relationship between (a) age 

of smoking initiation, any quit attempts, 

and a single nicotine dependence 

criterion, and (b) amount of lifetime 

smoking. Participants were given the 

Composite International Diagnostic 

Interview and the Fagerström Test for 

Nicotine Dependence.

The rates at which smokers who started 

smoking at a young age, had made five 

or more quit attempts, and met criteria 

for nicotine dependence increased with 

lifetime amount of smoking.

Jacobsen et al. 

(2005)

• 73 persons (41 

daily smokers and 

32 nonsmokers), 

14–18 years of age

• Exposure to nicotine (or 

not) was similar with 

respect to age, gender, 

and education level.

The study assessed several outcomes in 

daily smokers and nonsmokers: verbal 

working memory; verbal learning and 

memory; selected, divided, and sustained 

attention; mood; nicotine withdrawal; 

and tobacco craving. Analyses were 

controlled for general intelligence, 

reading achievement, parental education 

attainment, baseline affective symptoms, 

and lifetime exposure to alcohol and 

cannabis. 

In adolescent smokers, cessation of 

tobacco use increased tobacco craving, 

symptoms of withdrawal, and depressed 

mood. Adolescent smokers had 

impairments in accuracy of working 

memory regardless of how recently 

they had started smoking. Performance 

decrements were more severe with 

earlier age at onset of smoking. 

Adolescent smokers experienced 

further disruption of working memory 

and verbal memory during smoking 

cessation. Male smokers initiated 

smoking at earlier ages than females 

and were more impaired during tests 

of selective and divided attention than 

were female smokers and nonsmokers. 

Table A3.1-2 Continued
Evidence Tables A3.1-11

E-Cigarette Use Among Youth and Young Adults

Study Design/population Outcomes examined Findings

Lejuez et al. 

(2005)

• Cross-sectional analysis

• 125 students, 

predominantly African 

American, in grades 5–12

• Ever smokers (even a 

puff) and never smokers 

The relationship between propensity 

for risk taking and ever smoking 

was assessed using the standard and 

adolescent BART, as administered 

to ever and never smokers. The 

Sensation Seeking Scale and Eysenck 

Impulsiveness Scale were also 

administered. 

Ever smokers and never smokers 

differed on propensity for risk taking; 

higher BART scores and higher selfreported impulsive sensation-seeking 

scores were associated with greater odds 

of ever smoking. 

Biederman et al. 

(2006)

• Secondary analysis of 

a case-control family 

genetic study

• 300 females, 12 years of 

age and older, 97 with 

ADHD and 203 without 

ADHD 

• Females with ADHD 

were selected from 

referrals to a pediatric 

psychopharmacology 

clinic at a major 

academic center 

and from pediatric 

clinics of a major 

health maintenance 

organization.

• Controls were selected 

from the pediatric 

clinics.

Chronology of substance use was 

determined from participant recall 

of age at first use and age at which 

criteria for abuse or dependence were 

met. Survival models were used to 

assess whether cigarette smoking was 

associated with later alcohol and drug 

use and dependence. Results were 

adjusted for gender. 

A history of cigarette smoking was 

significantly associated with subsequent 

use of alcohol and drugs and with the 

development of alcohol and drug use 

disorders. The association was stronger 

in youth with ADHD than in controls.

Brook et al. 

(2007)

• Longitudinal cohort 

study

• Students in grades 7–10, 

of African American 

or Puerto Rican race/

ethnicity, in East 

Harlem, New York City

• Followed until they were 

in their mid-20s

Adolescents were assessed for tobacco, 

alcohol, and illicit drug use and 

dependence. Logistic regression was 

used to examine potential associations 

between patterns of cigarette use and 

subsequent alcohol and drug use and 

dependence. Results were controlled for 

ethnicity and gender.

Participants who had started smoking by 

age 14 and continued to smoke in their 

20s were more likely than nonsmokers, 

smokers who quit by their mid-20s, or 

smokers who started smoking between 

19 and 24 years of age to be diagnosed at 

follow-up with alcohol dependence and 

dependence on illicit drugs. Smokers 

who started between 19 and 24 years of 

age were more likely than nonsmokers 

to become drug but not alcoholdependent. 

DiFranza et al. 

(2007)

• Longitudinal cohort 

study

• 217 students in 6th grade 

who had inhaled tobacco 

smoke

• Six communities in 

Massachusetts

• Followed for 4 years

Outcomes included loss of autonomy 

as judged by using the HONC. Tobacco 

dependence was assessed using ICD-10 

criteria. Cox regression analysis was 

used to assess independent associations 

between potential predictors and 

outcomes. 

Fifty-eight percent of the students 

lost autonomy, and 38% developed 

ICD-10 dependence. Predictors of 

progression to loss of autonomy and 

tobacco dependence included a feeling 

of relaxation with the first cigarette and 

depressed mood. 

Table A3.1-2 Continued
A Report of the Surgeon General

A3.1-12 Appendix 3.1

Study Design/population Outcomes examined Findings

Jacobsen et al. 

(2007b)

• Cross-sectional analysis

• 67 adolescent smokers 

and nonsmokers, with 

and without prenatal 

exposure to maternal 

smoking

• Groups were similar with 

respect to age, education 

attainment, IQ, years of 

parental education, and 

symptoms of inattention. 

Diffusion tensor anisotropy and 

magnetic resonance imaging were 

used to examine white matter 

microstructure. The study assessed 

auditory attention and compared 

outcomes between exposed and 

unexposed participants.

Prenatal and adolescent exposure to 

tobacco smoke were both associated 

with increased fractional anisotropy 

in anterior cortical white matter. 

Adolescent smoking was associated 

with increased fractional anisotropy 

of regions of the internal capsule that 

contain auditory thalamocortical and 

corticofugal fibers. Fractional anisotropy 

of the posterior limb of the left internal 

capsule was positively correlated with 

reaction time during performance of 

an auditory attention task in smokers 

but not in nonsmokers. Development 

of anterior cortical and internal capsule 

fibers may be particularly vulnerable 

to disruption of cholinergic signaling 

induced by nicotine in tobacco smoke. 

Disruption of auditory corticofugal 

fibers may interfere with the ability 

of these fibers to modulate ascending 

auditory signals, leading to greater noise 

and reduced efficiency of neurocircuitry 

that support auditory processing.

Kandel et al. 

(2007)

• Longitudinal study

• 353 students in grades 

6–10 in public schools in 

Chicago

• One parent (usually the 

mother)

• Followed for 2 years 

Household interviews were conducted 

three times during the study period. 

Time to nicotine dependence criteria, 

per DSM-IV, and full dependence 

syndrome were assessed. Other potential 

risk factors assessed included race, 

gender, one or both parents ever being 

nicotine-dependent, positive diagnostic 

screen for conduct disorder, pleasant 

experience with first tobacco use, 

marijuana use before tobacco use, 

alcohol use before tobacco use, peer 

smoking, depressive symptoms, positive 

screen for anxiety disorder, academic 

performance, and pubertal stage. 

Twenty-five percent of the students 

experienced nicotine dependence 

within 23 months of onset of tobacco 

use. Tolerance, impaired control, and 

nicotine withdrawal were experienced 

most frequently. 

Musso et al. 

(2007)

• Cross-sectional analysis

• 27 White participants 

(15 regular smokers 

and 12 never smokers), 

18–25 years of age, who 

were recruited through a 

newspaper advertisement

Event-related functional magnetic 

resonance imaging was used to assess 

prefrontal attentional network function 

in young adults.

Prefrontal attentional network activity 

was significantly reduced in smokers 

compared with nonsmokers. Number 

of years of smoking was associated with 

the extent of diminished attentional 

network activity. 

Table A3.1-2 Continued
Evidence Tables A3.1-13

E-Cigarette Use Among Youth and Young Adults

Study Design/population Outcomes examined Findings

Bronisch et al. 

(2008)

• Longitudinal cohort 

study

• 3,021 adolescents and 

young adult participants, 

14–24 years of age at 

baseline, from the Early 

Development Stages of 

Psychopathology Study, 

randomly selected from 

population registers in 

Munich, Germany

• Followed for an average 

of 42 months

Participants were assessed for smoking 

behaviors, nicotine dependence, suicide 

ideation, and suicide attempts using 

the standardized Munich-Composite 

International Diagnostic Interview.

Occasional and regular smoking and 

nicotine dependence at baseline were 

strongly associated with suicide ideation 

and suicide attempts. Associations 

remained significant when participants 

with major depression were excluded. 

Wilens et al. 

(2008)

• Longitudinal cohort 

study

• 166 people (80 with 

ADHD and 86 control 

probands), 15–25 years of 

age, in Massachusetts

• Follow-up at 5 years for 

females and 10 years for 

males

This study assessed nicotine dependence 

in ADHD and control individuals. 

Nicotine dependence was assessed using 

the mFTQ. 

Smokers with ADHD had significantly 

higher scores on the mFTQ than 

smokers without ADHD. The study 

found a positive linear relationship 

between the mFTQ score and 

ADHD symptoms of inattention and 

hyperactivity. Parental smoking, peer 

smoking, and living with a smoker all 

increased the risk for smoking among 

those with ADHD compared with 

controls.

Boden et al. 

(2010)

• Longitudinal birth 

cohort study

• 1,055 children from the 

Christchurch Health 

and Development Study 

in Christchurch, New 

Zealand

• Followed from birth to 

25 years of age

This study assessed the relationships 

between smoking and depression in 

early adulthood using fixed-effects 

regression and structural equation 

modeling. Depression was assessed 

at 18, 21, and 25 years of age using a 

structured mental health interview. 

Smoking status and nicotine 

dependence also were assessed at 18, 21, 

and 25 years of age.

The study found significant associations 

between nicotine dependence symptoms 

and depressive symptoms. Findings 

remained significant after adjusting 

for potential confounders, including 

nonobserved genetic and environmental 

factors. Structural equation modeling 

suggested that the best-fitting causal 

model was one in which nicotine 

dependence led to depression. Use of 

daily cigarette intake, instead of nicotine 

dependence measures, produced similar 

findings.

Dierker and 

Mermelstein 

(2010)

• Longitudinal cohort 

study

• 694 students in grades 

9 and 10 from high 

schools in metropolitan 

Chicago: 594 students 

had smoked fewer 

than 100 cigarettes 

in their lifetime, and 

152 students had smoked 

100 or more cigarettes in 

their lifetime.

• Participants were drawn 

from the Social and 

Emotional Contexts of 

Adolescent Smoking 

Patterns Study. 

Nicotine dependence was assessed 

at baseline and at 24 months using 

the NDSS, modified for adolescents. 

Logistic regression was used to assess 

the association between nicotine 

dependence (determined by NDSS total 

score and endorsement of individual 

symptoms) at baseline and smoking 

behavior at 24 months. Results were 

adjusted for gender, number of days 

smoked in the past 30 days, number of 

cigarettes smoked in the past 7 days, 

and other tobacco use (chewing tobacco, 

snuff, or dip; cigars, cigarillos, or little 

cigars; and bidis or kreteks) in the past 

30 days. 

Among students who had smoked 

fewer than 100 cigarettes, high levels 

of nicotine dependence symptoms 

at baseline and individual symptoms 

predicted current and daily smoking 

behavior at 24-month follow-up; this 

finding was significant after controlling 

for baseline smoking (quantity and 

frequency), gender, and other tobacco 

use. Among students who had smoked 

100 or more cigarettes but were 

smoking fewer than 5 cigarettes/day, 

the level of nicotine dependence and 

individual symptoms did not predict 

smoking behavior at 24-month followup.

Table A3.1-2 Continued
A Report of the Surgeon General

A3.1-14 Appendix 3.1

Study Design/population Outcomes examined Findings

McKenzie et al. 

(2010) 

• Prospective cohort study 

of adolescent and young 

adult health

• Sample size = 1,943

• Teens were assessed 

6 times in monthly 

intervals, with two 

follow-up assessments 

during young adulthood 

(Wave 7, 1998; Wave 8, 

2001–2003)

Adolescent depression and anxiety 

symptoms were assessed using the 

Revised Clinical Interview Schedule 

(CIS-R). Young adult tobacco use was 

defined as daily use (6 or 7 days per 

week) and dependent use as ≥4 on 

the Fagerström Test for Nicotine 

Dependence).

Among adolescent “less than daily” 

smokers, those with high levels of 

depression and anxiety symptoms had an 

increased risk of nicotine dependence in 

young adulthood compared with young 

adults who had low levels of adolescent 

depression and anxiety symptoms, after 

adjusting for potential confounding 

factors.

Among adolescent “daily” smokers, 

those with high levels of depression 

and anxiety symptoms had an almost 

twofold increase in the odds of reporting 

nicotine dependence in young adulthood 

compared with young adults with low 

levels of adolescent depression and 

anxiety symptoms.

Griesler et al. 

(2011)

• Longitudinal cohort 

study

• 814 adolescent smokers 

in grades 6–10 and their 

mothers, as selected 

from the Transition to 

Nicotine Dependence 

study

• Adolescent participants 

were recruited from 

public schools in 

Chicago.

• Participants were 

followed for 2 years 

(five follow-ups for 

adolescents and three 

follow-ups for mothers).

Data from participants were obtained 

through computerized household 

interviews. Selected DSM-IV 

psychiatric disorders were ascertained 

annually from both adolescents and 

their mothers. Adolescent smoking, 

other tobacco use, DSM-IV nicotine 

dependence symptoms, and abuse and 

dependence on other substances were 

assessed. Maternal lifetime smoking, 

DSM-IV nicotine dependence symptoms, 

DSM-IV depression, and delinquency 

were also assessed. Cox proportional 

hazards models were used to estimate 

effects of (a) psychiatric disorders on 

the onset of nicotine dependence, and 

(b) nicotine dependence on psychiatric 

disorders. Multivariate models were used 

to control for psychiatric comorbidity 

and factors associated with nicotine 

dependence and psychiatric disorders.

Among lifetime smokers, 

53.7% experienced at least one nicotine 

dependence criterion; 26.1%, full 

dependence; 14.1%, an anxiety disorder; 

18.8%, a mood disorder; and 29.5%, a 

disruptive disorder. Nicotine-dependent 

youth had higher rates of individual 

and multiple disorders than those not 

dependent. After adjusting for covariates, 

mood disorder and nicotine dependence 

did not predict each other, but anxiety 

disorder predicted nicotine dependence. 

Bidirectional influences were observed 

for disruptive disorder and nicotine 

dependence. Predictors of full nicotine 

dependence included earlier age at onset 

of tobacco use; high initial pleasant 

sensitivity to tobacco; alcohol and illicit 

drug use, abuse, and dependence; and 

parental nicotine dependence. 

Jamal et al. 

(2011)

• Longitudinal cohort 

study

• 1,055 participants in the 

Netherlands Study of 

Depression and Anxiety 

Disorders who developed 

psychopathology after 

starting smoking 

Time to onset of psychopathology 

after starting smoking was assessed. 

Smoking behavior was measured by 

questionnaire; age at onset of smoking 

was defined as the age at which the 

participant began smoking regularly. 

Lifetime diagnosis and age at onset of 

depression and anxiety disorders were 

ascertained using the lifetime version of 

the Composite International Diagnostic 

Interview (version 2.1). Hierarchical 

multiple linear regression was used to 

examine the outcome of time between 

the onset of smoking and the onset of 

psychopathology. Results were adjusted 

for several potential confounders. 

The time between the onset of smoking 

and the onset of depression was 5 years 

shorter for early-onset than late-onset 

smokers. A greater percentage of earlyonset than late-onset smokers had the 

first onset of psychopathology within 

the first 5 years of starting smoking. 

Age at onset of smoking predicted the 

onset of psychopathology after adjusting 

for gender, education, and childhood 

trauma. When anxiety and depression 

disorder were examined separately, 

the pattern was true only for anxiety 

disorders. 

Table A3.1-2 Continued
Evidence Tables A3.1-15

E-Cigarette Use Among Youth and Young Adults

Study Design/population Outcomes examined Findings

Rubinstein et 

al. (2011)

• Cross-sectional analysis

• 12 adolescent smokers 

(1–5 cigarettes/day) 

and 12 nonsmokers, all 

13–17 years of age

Functional magnetic resonance imaging 

was used to assess responses to natural 

reinforcers (visual cues representing 

pleasurable foods).

Food images elicited greater activations 

in nonsmokers in multiple areas of the 

brain, including the insula, inferior 

frontal region, and rolandic operculum. 

Versus nonsmokers, smokers did not 

demonstrate greater blood oxygenationlevel-dependent activations in any 

region of the brain.

Cengelli et al. 

(2012)

• Systematic review of 

longitudinal, populationbased studies

• Adolescent and young 

adult smokers

Determinants of self-initiated smoking 

cessation lasting at least 6 months

Five factors robustly predicted quitting: 

not having friends who smoked, not 

having intention to smoke in the future, 

resisting peer pressure to smoke, being 

older at first use of cigarette, and having 

negative beliefs about smoking.

Moylan et al. 

(2012)

• Systematic review 

of population-based 

observational studies

• Random samples of 

adolescents and adults 

from Germany, the 

Netherlands, New 

Zealand, and the United 

States 

The studies in the review investigated 

associations between cigarette smoking, 

nicotine dependence, and anxiety 

disorders using recognized, structured 

clinical diagnostic criteria. 

Evidence suggests that some baseline 

anxiety disorders are risk factors 

for smoking initiation and nicotine 

dependence, but the evidence is 

heterogeneous. The evidence more 

consistently suggests that smoking and 

nicotine dependence are risk factors 

for the development of some anxiety 

disorders, such as panic disorder and 

generalized anxiety disorder, but the 

evidence is not consistent across studies. 

Dierker et al. 

(2012) 

• Face-to-face survey of 

adolescents (13–17 years 

of age)

• Sample size = 10,123

This study investigated the occurrence 

of nicotine dependence following the 

achievement of previous smoking 

milestones (initiation, weekly, and daily 

smoking).

Among adolescents who had ever 

smoked (36%), 40.7% had reached 

weekly smoking levels and 32.8% had 

reached daily smoking. Essentially one 

in five (19.6%) adolescents who had 

ever smoked met criteria for nicotine 

dependence. An earlier age of smoking 

initiation, a shorter time since the 

onset of smoking, and faster transitions 

between smoking milestones were 

independently associated with the 

onset of daily smoking and nicotine 

dependence.

Zhan et al. 

(2012) 

• Cohort study of 9th- and 

10th-grade students 

• Sample size = 12,970

This study investigated the natural 

course of nicotine dependence from 

first use/early initiation to actual 

dependence.

Nicotine dependence symptoms were 

reported before reaching 100 cigarettes 

for a substantial number of adolescents 

(from 9.4% to 58.8% for individual 

symptoms). 

Table A3.1-2 Continued
A Report of the Surgeon General

A3.1-16 Appendix 3.1

Study Design/population Outcomes examined Findings

Cavalca et al. 

(2013)

• Cross-sectional analysis

• 39 adolescents 

(22 smokers and 

17 nonsmokers) 

from high schools in 

Connecticut

• Smokers used five or 

more cigarettes per day.

• Nonsmokers reported 

never smoking in their 

lifetime. 

This study assessed associations 

between smoking status and risktaking behaviors. Smoking status was 

confirmed with urine cotinine level. 

Risk-taking behavior was measured 

using the standard BART and modified 

BART to include a peer component. 

Personality measures of impulsivity 

(assessed with the Barratt Impulsiveness 

Scale 11) and the degree to which 

subjects acted autonomously with peers 

(assessed with the Resistance to Peer 

Influence tool) were also measured. 

Smokers showed more risk-taking 

behaviors using the modified BART than 

with the standard BART. 

Galván et al. 

(2013)

• Cross-sectional analysis

• 18 daily smokers 

and 25 nonsmokers, 

17–21 years of age

This study compared brain function 

related to decision making in youth and 

young adult smokers and nonsmokers. 

The BART was administered to smokers 

and nonsmokers using functional 

magnetic resonance imaging to identify 

neural correlates of risky decision 

making.

Level of risk modulated the brain 

activities in the right dorsolateral and 

ventrolateral prefrontal cortices more in 

smokers than nonsmokers. 

Kendler et al. 

(2014)

• Monozygotic co-twin 

control study

• 175 male and 69 female 

twin pairs

• Virginia Adult Twin 

Study of Psychiatric and 

Substance Use Disorders 

Pairs were selected for discordant 

age at onset of regular smoking to 

study the association between age 

of onset of regular smoking and 

risk for later nicotine dependence. 

Nicotine dependence was assessed 

using the Fagerström Test for Nicotine 

Dependence and level of craving. Chisquare tests and t-tests were used to 

conduct comparisons. 

Twins who began smoking earlier 

had significantly higher scores on the 

Fagerström Test during their periods 

of heaviest smoking. Findings were 

consistent for male and female twin 

pairs. Craving for cigarettes when 

unable to smoke was also higher in the 

early-onset member of both groups. 

Early-onset twins did not differ from 

later-onset twins in symptoms of alcohol 

or cannabis use; maximal level of 

cannabis use; or sedative, stimulant, or 

cocaine use.

Tjora et al. 

(2014)

• Longitudinal cohort 

study

• 924 students in grade 7 

in the Norwegian 

Longitudinal Health 

Behaviour Study

• Followed from 13 to 

30 years of age

Cross-sectional and longitudinal 

analyses were conducted. The study 

measured daily smoking and depressed 

mood in nine waves. Depression and 

depressed mood were measured with 

an inventory developed by Alsaker 

(1992) and Holsen and colleagues 

(2000). Longitudinal models assessed 

depression predicting smoking, smoking 

predicting depression, and smoking and 

depression influencing each other from 

Waves 1 to 2 and Waves 2 to 3; the three 

models were compared with indices of 

fit. 

Cross-sectional analyses found an 

association between early adolescent 

depression and early adolescent 

smoking.

Longitudinal analyses showed early 

adolescent smoking predicting early 

adolescent depression and vice versa 

in Waves 1–2 and 2–3, and supporting 

reciprocal causal effects between 

smoking and depression that are 

established in early adolescence and 

maintained into adulthood.

Table A3.1-2 Continued
Evidence Tables A3.1-17

E-Cigarette Use Among Youth and Young Adults

Study Design/population Outcomes examined Findings

Treur et al. 

(2015)

• Cross-sectional and 

longitudinal study

• 1,987 adult and 

648 adolescent 

monozygotic twin 

pairs from the 

Netherlands Twin 

Registry, concordant and 

discordant for smoking

This study compared twins, in which 

pairs were concordant and discordant 

for tobacco exposure, for attention 

problems. Adult comparisons were 

cross-sectional, and adolescents were 

studied longitudinally from birth to 

approximately 18 years of age. 

Adult twins who had ever smoked had 

more attention problems than their 

never-smoking co-twins. Longitudinal 

analysis showed greater increases in 

attention problems from adolescence 

to adulthood in twins who ever smoked 

than in never-smoking co-twins. In 

childhood and adolescence, eversmoking twins had more attention 

problems than never-smoking co-twins; 

scores were similar before smoking was 

initiated and after both twins started 

smoking. 

Notes: ADHD = attention deficit hyperactivity disorder; BART = Balloon Analogue Risk Task; DSM-III = Diagnostic and Statistical 

Manual of Mental Disorders, 3rd Edition; DSM-IV = Diagnostic and Statistical Manual of Mental Disorders, 4th Edition; 

HONC = Hooked on Nicotine Checklist; ICD-10 = International Statistical Classification of Diseases and Related Health Problems, 

10th revision; IQ = intelligence quotient; mFTQ = modified Fagerström Tolerance Questionnaire; NDSS = Nicotine Dependence 

Syndrome Scale; NHANES III = Third National Health and Nutrition Examination Survey; NHSDA = National Household Survey on 

Drug Abuse.

Table A3.1-2 Continued
A Report of the Surgeon General

A3.1-18 Appendix 3.1

Table A3.1-3 Preclinical/animal studies on adolescent nicotine exposure

Study Species/exposure Measures Outcome/findings Comments

Williams and 

Kanagasabai (1984) 

• Sprague-Dawley rats were 

mated and assigned to control 

and nicotine groups.

• Animals were weighed at 

regular intervals and killed on 

day 20 of pregnancy.

• Fetal and placental weights were 

recorded, and analysis of fetal 

body water, fat, protein, and 

DNA carried out.

• Rates of maternal adipose tissue 

lipolysis and lipogenesis were 

measured.

• Weight gains of mothers in 

the nicotine group were lower 

in the first and second weeks 

of pregnancy but similar to 

controls in the third week. 

• Fetal body weight, DNA, protein, 

and percentage water content 

were similar in the two groups. 

Mean fetal body fat (g/kg) was 

significantly higher in the 

nicotine group (96.2) than in 

controls (72.0). 

• Rates of maternal lipolysis were 

also higher in the nicotine 

group.

—

Newman et al. (1999) • Rat offspring were exposed to 

nicotine through implantation 

of osmotic minipumps in 

dams at levels of 0.75, 1.5, 

and 3.0 mg/kg/day for 19 days 

prenatally and 16 days 

postnatally.

• Gestation length, body weight, 

litter size, sex difference, and 

locomotor activity.

• No significant effects were 

shown for gestation length, 

litter size, or male-to-female 

pup ratio.

• Higher percentage of pup deaths 

resulted from nicotine-exposed 

dams than from control dams.

• Significantly less litter body 

weight was shown in nicotineexposed offspring on P1 when 

compared with controls. 

However, these offspring 

surpassed the control groups in 

litter body weight on P14 and 

P21.

• Hyperactivity was shown in 

offspring exposed to prenatal/

postnatal nicotine at levels 

of 0.75 and 3.0 mg/kg/day on 

P14, but not on P21 or at the 

1.5 mg/kg/day condition.

Results are consistent with the 

hypothesis that rat offspring are 

susceptible to the neurochemical 

and neurobehavioral effects 

of prenatal/postnatal nicotine 

exposure.
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-19

Study Species/exposure Measures Outcome/findings Comments

Grove et al. (2001) • Pregnant rhesus monkeys were 

treated with nicotine tartrate 

(1.5 mg/kg x d) starting on 

day 26 of pregnancy and then 

through day 160 of gestation.

• Weight of offspring at birth 

• P1 plasma leptin level

• Nicotine exposure had no 

significant effect on absolute 

birth weights of the neonatal 

monkeys, although there 

was a 10% reduction in birth 

weights with nicotine exposure 

when they were normalized to 

maternal weight.

• P1 plasma leptin levels were 

significantly reduced by about 

50% in the nicotine treatment 

group compared with saline 

controls, suggesting that the 

infant monkeys exposed to 

nicotine may also have had 

lower body fat levels.

Findings from this study suggest 

that nicotine exposure during 

pregnancy may increase energy 

expenditure in the developing 

fetus through actions on 

hypothalamic systems, resulting 

in lower birth weights and body 

fat levels.

Adriani et al. (2002) • Outbred CD-1 mice in early 

(P28–P35), middle (P37–48), 

and late (P50–P61) adolescence

• Oral self-administration 

(10 mg/L nicotine)

• Reinforcing effects of nicotine—

oral self-administration with 

two-bottle choice (addiction/

rewarding)

• Early adolescents showed 

significant preference for 

nicotine-containing solution 

(intake was 1.15 ± 0.04 mg/kg 

on final day); middle adolescents 

had no preference; late 

adolescents showed aversion to 

nicotine.

Early adolescents (P28−P35) 

were the only group to increase 

consumption when nicotine 

content was lowered.

Vastola et al. (2002) • Male and female SpragueDawley rats

• Conditioned with saline or 

nicotine (weight of salt was 

0.6 mg/kg, s.c.) was begun on 

P28 or P58.

• Testing occurred on P40 or 

P70, respectively, in a biased 

CPP paradigm.

• Rewarding effects of nicotine

• CPP

• Adolescent rats of both genders 

showed a significant place 

preference for nicotine, while 

adults did not. 

• Nicotine-induced locomotion 

during CPP testing also showed 

age effects. 

• Adults were sensitive to motorsuppressing effects of nicotine, 

while adolescents showed some 

sensitization after multiple 

exposures.

Adolescents were more sensitive 

than adults to the rewarding and 

locomotor-activating effects of 

nicotine. A possible mechanism 

for this age difference is an 

increase in NAc dopamine tone 

across development that leads 

to decreased stimulation of 

mesolimbic structures and less 

sensitivity to nicotine.

Table A3.1-3 Continued
A Report of the Surgeon General

A3.1-20 Appendix 3.1

Study Species/exposure Measures Outcome/findings Comments

Levin et al. (2003) • Female Sprague-Dawley 

rats (~P54 or ~P84 start); 

acute and chronic nicotine 

i.v. self-administration; 

0.01–0.8 mg/kg/infusion 

for acute-dose response; 

0.03 mg/kg/infusion for chronic 

testing

• Reinforcing effects of nicotine—

adult vs. adolescent onset of 

acute and chronic i.v. selfadministration 

• Adolescents had almost double 

the rate of self-administration 

as adults at 0.03 mg/kg/infusion 

during chronic testing.

Adolescent onset of selfadministration resulted in greater 

nicotine intake, and this effect 

persisted into adulthood.

Slawecki et al. (2003) • Sprague-Dawley rats

• Transdermal nicotine patches 

delivered 5 mg/kg/day nicotine 

from P31 to P36.

• Testing occurred at least 10 days 

after nicotine exposure.

• Emotional responding in 

adulthood

• Nicotine-exposed rats had lower 

ambulatory counts, centersquare crosses, and latency to 

retreat than nonexposed rats.

• In a modified open-field test 

with food in the center, exposed 

rats approached food and spent 

significantly less time with food 

than unexposed rats.

Adolescent nicotine produced 

lasting increases in anxiety-like 

behavior, even when a food reward 

was available. The mechanism 

may be enhanced CRF signaling in 

the amygdala.

Belluzzi et al. (2004) • Sprague-Dawley rats at P28, 

P38, or P90

• Single-trial CPP with 

0.125–0.5 mg/kg nicotine

• Rewarding effects of nicotine

• Single-trial CPP (addiction/

rewarding)

• Early adolescents (P28) showed 

CPP for 0.5 mg/kg nicotine, a 

subthreshold dose. 

Early adolescents showed greater 

sensitivity to the rewarding effects 

of a single nicotine injection 

(0.5 mg/kg). Adults and late 

adolescents (P38) did not develop 

place preference for any dose.

Schochet et al. 

(2005)

• Male Sprague-Dawley rats

• Rats received acute injections 

of saline or nicotine (0.1 or 

0.4 mg/kg, s.c.) at 30 days of age 

or 70 days of age.

• Nicotine-induced neural 

activation

• Age-related differences were 

found in baseline levels of arc 

and cFos, with adolescents 

having higher expression than 

adults. 

• Nicotine produced greater arc 

induction in adolescent PFC 

regions than adults, especially 

in the ventrolateral orbital 

cortex and somatosensory 

cortex.

• cFos and NGFI-B were 

upregulated by nicotine in an 

age-independent manner.

Adolescents may have higher 

baseline neuronal activity, and 

nicotine exposure during this 

time may have more profound 

activational effects in regions 

mediating reward and executive 

function than in adulthood. This 

may help explain age-related 

behavioral differences seen in 

responses to nicotine.

Table A3.1-3 Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-21

Study Species/exposure Measures Outcome/findings Comments

Chen and Kelly 

(2005) 

• Pregnant Sprague-Dawley rats

• Rats were given 0-, 15-, 

or 25-mg nicotine pellets 

throughout pregnancy.

• One group of offspring received 

1 or 2 mg/kg/day nicotine 

during early postnatal period.

• One group of offspring received 

no nicotine. 

• Body weight of offspring

• Neonatal thyroid status

• Nicotine treatment significantly 

increased body weight in female 

offspring starting on P35 and 

continued into adulthood. 

• This increase was seen initially 

in male offspring but did not 

persist into adulthood. 

Findings support other studies 

reporting a higher body weight 

among children born to mothers 

who smoked during pregnancy. 

Data from this study on thyroid 

status suggest that cigarette 

smoking-induced alterations in 

thyroid status might be mediated 

through compounds in cigarettes 

other than nicotine. 

Cohen et al. (2005) • Offspring of Wistar and 

mutant mice lacking beta2 and 

nAChR subunit

• Pregnant mice were implanted 

with osmotic minipumps that 

delivered either water or a 

controlled dose of nicotine.

• Sympathoadrenal system 

development in offspring 

• Breathing and arousal reflexes 

in offspring 

• Newborn Wistar pups exposed 

to nicotine exhibited all of 

the deficits associated with 

maternal tobacco and nicotine 

use and linked to poor 

neonatal outcome: growth 

restriction, unstable breathing, 

and impaired arousal and 

catecholamine biosynthesis.

• Similar deficits were detected 

in offspring lacking beta2-

containing nAChRs.

Article suggests that the 

underlying mechanisms of 

nicotine’s detrimental side effects 

on a range of crucial defensive 

reflexes involve loss of function 

of nAChR subtypes, possibly via 

activity-dependent desensitization.

Holloway et al. 

(2005) 

• Offspring of female Wistar 

rats were given either 

saline (vehicle) or nicotine 

(1 mg/kg-1 day-1) during 

pregnancy and lactation.

• Effect of fetal and neonatal 

exposure to nicotine, the 

major addictive component 

of cigarettes, on postnatal 

growth, adiposity, and glucose 

homeostasis

• Exposure to nicotine resulted in 

increased postnatal growth and 

adiposity. Nicotine exposure also 

resulted in dysglycemia at 7 and 

26 weeks of age. Serum insulin 

concentrations were decreased 

in the pups exposed to nicotine 

at birth. This was associated 

with increased beta cell 

apoptosis (pups of saline-treated 

mothers, 8.8 (±1.21% apoptotic 

beta cells; pups of nicotinetreated mothers, 27.8 (±3.1%) 

apoptotic beta cells).

Fetal and neonatal exposure to 

nicotine results in metabolic 

changes in the offspring that are 

consistent with obesity and type 2 

diabetes. Authors proposed that 

observed metabolic changes may 

be a consequence of the initial 

insult to the beta cell during fetal 

life and that this animal model has 

many characteristics of diabetes in 

humans.

Table A3.1-3 Continued
A Report of the Surgeon General

A3.1-22 Appendix 3.1

Study Species/exposure Measures Outcome/findings Comments

Gao et al. (2005) • Offspring of Wistar rats were 

given nicotine or saline during 

pregnancy and lactation.

• Postnatal growth in offspring 

from weaning to 26 weeks

• Exposure to nicotine resulted 

in increased postnatal body 

weight and fat pad weight 

and an increased amount of 

perivascular adipose tissue 

(PVAT) in the offspring.

• Phenylephrine-induced 

contraction without PVAT was 

not different between salineand nicotine-exposed rats.

Prenatal nicotine exposure 

increased adiposity and caused 

an alteration in the modulatory 

function of PVAT on vascular 

relaxation response.

O’Dell et al. (2006) • Male Wistar rats

• Osmotic minipumps 

delivered saline or nicotine 

(0-4.7 mg/kg/day free base or 

3.2 mg/kg/day free base) from 

~P29 to P36 for adolescents and 

from ~P36 to P44 for adults to 

compare somatic and affective 

withdrawal.

• Withdrawal was precipitated 

in both groups with 

mecamylamine. 

• Somatic and affective 

withdrawal from chronic 

nicotine

• Adults displayed elevated ICSS 

thresholds during precipitated 

withdrawal, while adolescents 

showed no change. 

• Adults also displayed a dosedependent increase in somatic 

withdrawal symptoms, while 

mecamylamine precipitated 

significant withdrawal in 

adolescents only at the highest 

dose (3.0 mg/kg, i.p.).

Overall, adolescents showed 

decreased sensitivity to the 

somatic and negative affective 

symptoms of nicotine withdrawal 

versus adults. These effects are 

not likely because of metabolic 

differences because blood 

nicotine levels were similar. The 

mechanism may be immature 

cholinergic and/or dopaminergic 

neurotransmission in the VTA. 

Shram et al. (2006) • Male Wistar rats

• 10-day CPP with saline or 

nicotine (0.2, 0.4, or 0.8 mg/kg 

free base, s.c.) conditioning 

followed by testing on P38 or 

P70–P73

• 4-day CTA with saccharin 

and nicotine (0, 0.2, 0.4, or 

0.8 mg/kg free base, s.c.) 

pairings followed by access to 

water or saccharin on P35 or 

P70–P73 

• Rewarding and aversive 

properties of nicotine

• CPP and CTA

• Adolescent rats displayed CPP 

for 0.8 mg/kg nicotine, while 

adults did not display CPP for 

any dose. 

• In CTA, nicotine pairings in 

adults attenuated saccharin 

consumption at the 0.4-mg/kg 

dose and trends for attenuation 

at 0.2 and 0.8 mg/kg.

• Adolescents did not show any 

change in saccharin intake.

Periadolescent rats were less 

sensitive to the aversive effects 

of nicotine; results suggest that 

reward-aversion balance may shift 

toward reward in adolescents. 

Two possible substrates proposed: 

immature mesolimbic dopamine 

system and immature cholinergic 

system.

Table A3.1-3 Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-23

Study Species/exposure Measures Outcome/findings Comments

Smith et al. (2006) • Male and female Long- Evans 

rats

• Osmotic minipumps 

delivered saline or nicotine 

(1 or 2 mg/kg/day free base) 

from P28 to P42 or P85 to P99.

• Testing was done 1 month after 

treatment.

• Emotional responding in 

adulthood

• High- and low-dose-exposed 

adolescents spent less time in 

the center of the open-field 

arena.

• Low-dose exposure to nicotine 

enhanced fear conditioning. 

However, extinction of freezing 

behavior was impaired after lowdose exposure. 

• Adult exposed rats overall 

displayed greater freezing in 

cued-fear conditioning than 

adolescent exposed animals, 

regardless of treatment.

• No gender-related differences 

were seen.

Adolescent nicotine produced 

lasting increases in anxiety-like 

behavior and enhanced fear 

learning with impaired extinction. 

This may be because of increased 

α4 nAChR mRNA levels in PFC.

Brielmaier et al. 

(2007)

• Male Sprague-Dawley rats

• Single-trial place conditioning 

began on P28 or P77 with saline 

or nicotine (0.5 mg/kg free 

base, s.c.), and testing occurred 

2 days later.

• Rewarding effects of nicotine

• CPP

• Adolescents developed a 

significant place preference for a 

single nicotine exposure, while 

adults did not. 

Age-related differences in 

sensitivity to nicotine were 

likely attributable to continuing 

maturation of cholinergic or 

mesocorticolimbic dopamine 

systems.

Britton et al. (2007) • Pregnant Sprague-Dawley rats

• Osmotic minipumps delivered 

saline or nicotine (2 mg/kg/day 

free base) from gestational day 7 

to pup weaning on P21. 

• nAChR signaling in adolescence

• Effects of perinatal nicotine 

exposure were measured using 

nicotine-stimulated 86Rb+ 

efflux assay in the frontal cortex, 

hippocampus, striatum, and 

thalamus.

• Cholinergic signaling via 

nAChRs was assessed in 

adolescent (P28, P35, and P49) 

and adult (P69) offspring. 

• In controls, nicotine-stimulated 

86Rb+ peaked at midadolescence (P35 and/or P49) in 

all regions.

• Perinatal nicotine eliminated 

the peak.

nAChR function, especially 

of the α4β2 subtype, peaked 

during adolescence. This may be 

attributable to higher expression, 

or differences in desensitization 

and recovery from desensitization. 

Perinatal nicotine prevented 

normal developmental trajectory 

of nAChR function, possibly 

attributable to decreased receptor 

expression or binding. This 

contrasts with clinical work 

showing greater sensitivity to 

nicotine in adolescence.

Table A3.1-3 Continued
A Report of the Surgeon General

A3.1-24 Appendix 3.1

Study Species/exposure Measures Outcome/findings Comments

Chen et al. (2007) • Lewis rats, P43−P45 to 

P55−P57; 10-day, prolonged 

access (23 hours/day) i.v. 

self-administration of saline 

or nicotine (0, 7.5, 15, 30, or 

60 μg/kg/infusion, free base)

• Reinforcing effects of i.v. 

nicotine

• Intravenous self-administration 

with prolonged access

• Male and female adolescents 

acquired self-administration 

of nicotine at 15 and 30 μg/kg 

doses but did not acquire the 

behavior at 7.5 μg/kg dose.

• Age comparison in females 

at 30 μg/kg dose showed that 

adolescents self-administered 

significantly more nicotine and 

acquired the behavior more 

rapidly than adults.

Observed no gender-based 

differences in sensitivity to 

nicotine dose in adolescent rats, 

with both genders showing an 

inverted U-shaped dose–response 

relationship, although rates of 

self-administration were still high 

at the higher doses. Adult females 

showed slower acquisition and 

decreased intake than adolescent 

females. Age differences may 

be attributable to enhanced 

metabolism, learning, or druginduced corticosterone release in 

adolescents. 

Kota et al. (2007) • Adolescent (P28–P36) and adult 

(P75–P84) ICR mice

• CPP (0.05–1 mg/kg, s.c. 

injections of nicotine or saline)

• Withdrawal (osmotic minipumps 

delivered saline or nicotine 

[48 mg/kg/day]) for 7 days; 

on day 8, minipumps were 

removed to assess spontaneous 

withdrawal, or withdrawal was 

precipitated with 2.0 mg/kg, 

s.c. mecamylamine.

• Anxiety-like and somatic 

withdrawal symptoms were 

scored.

• nAChR signaling in adolescence 

and adulthood

• CPP, nicotine-stimulated 

86Rb+ efflux, spontaneous and 

precipitated withdrawal 

• Adolescents had increased 

sensitivity (versus adults) to 

rewarding effects of low nicotine 

doses (0.05 and 0.1 mg/kg) in a 

CPP paradigm.

• In both spontaneous and 

precipitated withdrawal, 

adolescents showed fewer 

somatic and affective symptoms 

than adults.

• nAChR function was potentiated 

in adolescents, who showed a 

leftward shift of the nicotine 

dose–response curve in the 

86Rb+ efflux assay. 

Adolescents were more sensitive 

to the rewarding effects of 

nicotine, but they exhibited 

fewer withdrawal symptoms 

than adults during precipitated 

and spontaneous withdrawal. A 

possible mechanism is increased 

function of nAChRs, also found 

here. Increased nAChR expression 

may also play a role, but it was not 

tested herein.

Table A3.1-3 Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-25

Study Species/exposure Measures Outcome/findings Comments

McQuown et al. 

(2007)

• Male and female SpragueDawley rats (P28–P32 or 

P86–P90)

• Rats were pretreated with 

nicotine (60 μg/kg/day, i.v.) for 

4 days; with sucrose or cocaine 

(200 or 500 μg/kg/infusion, 

i.v.) self-administration the 

following 5 days.

• Reinforcing effects of other 

drugs of abuse

• Acquisition of cocaine selfadministration

• Adolescents had greater cocaine 

intake on all days of selfadministration than adults.

• Sucrose self-administration was 

unaffected by pretreatment.

Low-dose, semichronic nicotine 

exposure in early adolescence 

enhanced the reinforcing 

properties of cocaine but 

not sucrose. This difference 

is potentially attributable to 

immaturity of corticolimbic 

dopamine systems. This result 

suggests a “gateway” effect of 

adolescent nicotine on drug 

rewards but not natural rewards.

Shram et al. (2007) • Male Wistar rats

• Rats received acute injections 

of saline or nicotine (0.4 or 

0.8 mg/kg, s.c.) on P35 or 

P67–P70.

• Nicotine-induced neural 

activation

• Adolescents had higher basal 

cFos levels in paraventricular 

nucleus of the thalamus, medial 

septum, and ventral tegmental 

area, while adults had higher 

cFos in pedunculopontine 

tegmental nucleus.

• Nicotine-induced cFos in 

multiple subcortical regions, 

with adolescents showing 

greater induction in BNST, PVN 

of the thalamus, VTA, and NAcshell.

Regions important for reward 

processing, such as the VTA, 

NAc-shell, and BNST, were more 

sensitive to nicotine-induced 

cFos increases in adolescent rats 

than in adult rats. Immature 

cholinergic and dopaminergic 

systems or different nicotine 

pharmacokinetics likely underlie 

age-dependent responses to acute 

nicotine.

Bergstrom et al. 

(2008)

• Long-Evans rats

• Osmotic minipumps delivered 

saline or nicotine (2 mg/kg/day) 

from P29 to P43 or P80 to P93.

• Tissue was collected 5 weeks 

later for Golgi-Cox staining.

• Synaptic remodeling

• Layer V pyramidal neurons

• Nicotine exposure during 

adulthood increased basilar 

dendritic length and branch 

number in simple neurons.

• Adolescent nicotine exposure 

led to a modest increase in 

basilar dendritic length in 

complex neurons.

Both adolescent and adult nicotine 

exposure altered pyramidal 

neuron structure but in distinct 

morphologic subtypes depending 

on age. 

Table A3.1-3 Continued
A Report of the Surgeon General

A3.1-26 Appendix 3.1

Study Species/exposure Measures Outcome/findings Comments

Doura et al. (2008) • Sprague-Dawley rats

• Osmotic minipumps delivered 

saline or nicotine (6 mg/kg/day 

free base) for 14 days starting on 

P29 or in adulthood (~P70–P90)

• Tissue collected on day 14.

• nAChR expression

• Quantitative autoradiography 

for measurement of α7 

α4β2* and α6* nAChRs using 

[125I]αBtx, [125I]A-85380, 

and [125I]α-Ctx MII binding, 

respectively

• In saline-treated adolescents, 

binding at a4b2 nAChRs was 

uniformly higher across the 

38 regions analyzed than in 

saline-treated adults, with a 

50% mean increase.

• Nicotine-induced upregulation 

of a4b2* nAChRs was less 

profound in adolescents.

• Saline-treated adolescents had 

higher binding at a7, while 

nicotine treatment induced less 

upregulation than in adults.

•	 a6 binding was largely similar 

across ages in saline-treated 

animals, but adolescents showed 

downregulation after nicotine, 

while adults showed no change 

in a6 binding.

α4β2* and α7 nAChRs showed 

a developmental peak during 

adolescence and a decreased 

sensitivity to nicotine-induced 

upregulation. This age difference 

in saline-treated animals may 

be explained by normal synaptic 

pruning in adolescence. α6* 

nAChRs, on the other hand, do 

not show a similar developmental 

arc and nicotine-induced 

downregulation that is not seen in 

adults. This may be attributable to 

unique subunit compositions in 

α6* nAChRs in adolescents. 

Torres et al. (2008) • Male Wistar rats

• 10-day CPP with saline, nicotine 

(0–1.8 mg/kg free base, s.c.), or 

damphetamine (0–2.0 mg/kg 

salt, s.c.) conditioning followed 

by testing on P42 or P74. 

Adult rats were drug-naïve or 

pre-exposed to nicotine via 

osmotic minipumps delivering 

4.7 mg/kg/day from P28 to P42.

• Rewarding and aversive 

properties of nicotine

• CPP

• Naïve rats developed preference 

for nicotine in an inverted 

U-shape dose response, 

although adolescents developed 

preference for a wider range of 

doses than adults.

• Adults developed preference for 

0.2 mg/kg nicotine only.

• In pre-exposed adults, no 

preference developed for the 

0.2 mg/kg dose, but aversion 

to the 1.2 mg/kg dose did not 

develop.

• Adolescents also showed 

diminished aversion to high doses 

(1.2 or 1.8 mg/kg). Conditioning 

with d-amphetamine showed no 

age differences in development 

of CPP. 

Adolescents were more sensitive 

to the rewarding properties and 

less sensitive to the aversive 

properties of nicotine, but 

they showed no age differences 

in sensitivity to the effects of 

d-amphetamine. Pre-exposure 

to nicotine during adolescence 

blocked CPP in adulthood without 

causing tolerance and decreased 

aversion to high nicotine doses.

Table A3.1-3 Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-27

Study Species/exposure Measures Outcome/findings Comments

Counotte et al. 

(2009)

• Male Wistar rats

• Thrice-daily saline or nicotine 

(0.4 mg/kg, s.c.) from P34 to 

P43 or P60 to P69

• Behavioral testing occurred 

5 weeks later.

• Attention/cognition

• 5-CSRTT or delayed-reward task

• In 5-CSRTT, adolescent nicotine 

had detrimental effects on 

accuracy (i.e., reduction in 

correct detection) and increased 

impulsivity and time-out 

responses.

• There were no effects of 

treatment on the delayedreward task.

Adolescent exposure to nicotine 

produced lasting deficits in 

visuospatial attention and 

increased impulsive action but not 

impulsive choice. The mechanism 

may be hyperresponsiveness of 

dopaminergic terminals in medial 

PFC or alterations of serotonergic 

signaling.

Iñiguez et al. (2009) • Male Sprague-Dawley rats (P30)

• Two injections of nicotine 

(0.32 mg/kg, s.c.) on P30

• Behavioral testing on P85

• Mood

• FST (sensitivity to aversive 

stimuli)

• During FST in adulthood, 

nicotine exposure produced 

higher immobility and lower 

swimming counts than in 

vehicle-treated rats.

Single nicotine exposure during 

adolescence enhanced sensitivity 

to aversive stimuli in adulthood. 

Chronic nicotine’s ability to 

produce persistent depressivelike symptoms was blocked by 

acute nicotine or antidepressant 

treatment.

Counotte et al. 

(2011)

• Wistar rats (P34–P43 or 

P60–P69 during drug 

treatment)

• Thrice-daily saline or nicotine 

(0.4 mg/kg, s.c.) for 10 days

• Behavioral testing 5 weeks later

• Attention, cognition

• 5-CSRTT 

• Measured 5-CSRTT and synaptic 

protein expression and function 

in adulthood.

• More impulsivity, impaired 

attention, and downregulation 

of mGluR2 in PFC of 

adolescent-treated rats only.

Adolescent, but not postadolescent, 

nicotine treatment led to 

diminished attention span and 

greater impulsivity as adults. 

Deficits were associated with 

reduction of presynaptic mGluR2 

expression and function in PFC; 

group II mGluR2 agonist rescues 

deficits.

Table A3.1-3 Continued
A Report of the Surgeon General

A3.1-28 Appendix 3.1

Study Species/exposure Measures Outcome/findings Comments

Dao et al. (2011) • Male Sprague-Dawley rats

• Rats were treated with nicotine 

(60 μg/kg/day, i.v.) for 1 or 

4 days starting at P28 or P86.

• Nicotine-induced neural 

activation

• Acute nicotine increased cFos

in the NAc-shell and BLA of 

adolescents but not adults.

• Subchronic nicotine induced 

significant cFos expression 

in the BLA of both ages but a 

greater increase in the NAcshell 

of adolescents.

• Basal cFos differences were seen 

in PFC, NAc core, and dorsal 

striatum that were unaffected by 

acute or subchronic nicotine. 

Acute or subchronic nicotine 

during adolescence had more 

profound activational effects 

in regions mediating reward 

and executive function than in 

adulthood. This may help to 

explain age-related behavioral 

differences seen in responses 

to nicotine, such as increased 

cocaine self-administration and 

quinpirole-induced locomotion.

Goriounova and 

Mansvelder (2012)

• Male Wistar rats

• Thrice-daily injections of saline 

or nicotine (0.4 mg/kg free base, 

s.c.) from P34 to P43 or P60 to 

P69

• Tissue collected for 

electrophysiology

1–4 days or 5 weeks after 

treatment.

• Spike-timing-dependent 

plasticity in the PFC

• In adolescent control cells, 

induction of tLTP resulted in 

increased synaptic strength of 

130% ± 15% for 45 minutes in 

the medial PFC.

• This increase was not seen 

after acute bath application of 

nicotine. 

• Four days after adolescent 

nicotine exposure, tLTP 

induction resulted in less 

potentiation than in salinetreated animals. 

• Five weeks after nicotine 

treatment, exposed adults 

showed greater potentiation 

after a single action potential 

tLTP induction protocol than 

saline-treated adults, but no 

change in potentiation to a 

5 AP tLTP induction protocol. 

• Nicotine treatment in 

adulthood had no effect on 

spike-timing-dependent 

plasticity. 

Adolescent nicotine exposure 

produced lasting alterations 

in excitatory synapses and a 

resulting increase in tLTP. In 

the short term, reduced tLTP 

induction may be attributable 

to nicotine-induced increases in 

mGluR2 levels, while persistent 

reductions in inhibitory mGluR2 

signaling from nicotine allowed 

for increased tLTP in adults.

Table A3.1-3 Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-29

Study Species/exposure Measures Outcome/findings Comments

Portugal et al. (2012) • Male C57BL/6J mice

• Mice received acute nicotine 

(0, 0.045, 0.09, or 0.18 mg/kg 

free base, i.p., 5 minutes before 

training) or chronic nicotine 

via osmotic minipumps (0, 3.0, 

6.3, or 12 mg/kg free base) for 

12 days starting at P23, P38, or 

P53.

• Testing occurred 24 hours 

later (chronic nicotine); 

24 hours after pump removal 

(withdrawal); or 30 days after 

pump removal (prior chronic 

nicotine).

• Learning and memory • Acute nicotine enhanced 

contextual fear conditioning in 

all ages, although this occurred 

across a broader range of doses 

in preadolescents.

• Withdrawal impaired contextual 

fear conditioning in all ages, but 

at different doses.

• Additional testing found that 

adolescent mice withdrawing 

from 1.1 mg/kg/day also showed 

impairments.

• Prior exposure to nicotine 

during preadolescence or 

adolescence impaired contextual 

fear conditioning in adulthood, 

but adult nicotine had no effect.

• No changes in cued-fear 

conditioning were seen with any 

age or treatment.

Adolescents were especially 

sensitive to nicotine withdrawalinduced deficits in hippocampaldependent contextual fear 

conditioning, showing 

impairments across a wide range 

of doses. The mechanism may be 

lack of drug-induced upregulation 

of high-affinity nAChR binding 

in the hippocampus, decreases 

in total hippocampal CREB, 

or differences in nAChR 

desensitization. 

Natividad et al. 

(2013)

• Wistar rats

• Drug-naïve adolescents and 

adults or adolescent nicotineexposed adults began prolonged 

access (23 hours/day) i.v. selfadministration of saline or 

increasing nicotine (0.03, 0.06, 

or 0.09 mg/kg/0.1 mL 

infusion) at P43–P45 or P75 

in a 4 days-on and 3 days-off 

extinction paradigm.

• Exposed adolescents had 

minipumps deliver nicotine 

(4.7 mg/kg/day) for 14 days.

• Reinforcing effects of nicotine

• Intravenous self-administration 

with prolonged access and 

periods of forced abstinence

• Adolescents had the highest 

levels of nicotine intake at all 

doses, an increase in responding 

after each period of forced 

abstinence, and the highest 

responding during extinction. 

• Pre-exposed adults had the 

second-highest nicotine intake, 

followed by naïve adults.

• Adolescents also showed high 

nonreinforced responses relative 

to other groups.

Adolescents seemed to be 

particularly sensitive to the 

reinforcing or motivational 

properties of nicotine, and 

exposure during this period 

produced lasting increases 

in nicotine reward. High 

nonreinforced responding in 

adolescents may reflect increased 

impulsivity, motivation, or druginduced activity.

Table A3.1-3 Continued
A Report of the Surgeon General

A3.1-30 Appendix 3.1

Study Species/exposure Measures Outcome/findings Comments

Pickens et al. (2013) • Male and female Long- Evans 

rats

• Daily saline or nicotine 

(1 mg/kg, i.p.) injections from 

P25 to P59

• 35-day drug-free period before 

7 weeks of SMC testing

• Attention/cognition

• SMC task

• Regardless of treatment, females 

learned chunk-boundary and 

violation elements more slowly 

than males.

• Nicotine exposure impaired 

chunk-boundary element 

learning in males only, but it 

impaired violation elements in 

females. 

• Extra training abolished 

drug-induced impairments in 

learning.

Adolescent nicotine exposure 

produced lasting impairments in 

stimulus–response discrimination, 

but not abstract rule learning, in 

a gender-dependent manner. The 

mechanism may be impairment 

of hippocampal-dependent 

memory systems or drug-induced 

alterations in nAChR expression 

or function in hippocampal 

circuits.

Dickson et al. (2014) • C57BL/6J mice

• Osmotic minipumps delivered 

saline or nicotine (24 mg/kg/day 

free base) from P28 to P56.

• 5-day cocaine self-administration 

(1.0 mg/kg, i.v.) from P73 to P77 

before dose–response analysis 

with 0.18–1.8 mg/kg cocaine

• Reinforcing effects of other 

drugs of abuse

• Acquisition of cocaine selfadministration

• All animals acquired cocaine 

self-administration behavior at 

the 1.0 mg/kg dose.

• Adolescent nicotine exposure 

had no effect.

• In a dose–response analysis, 

adolescent nicotine exposure 

significantly increased cocaine 

intake at all but the highest 

1.8 mg/kg dose, compared with 

unexposed adults.

Greater cocaine intake in exposed 

animals during dose–response 

may be attributable to nicotineinduced hyperactivity, in contrast 

to McQuown and colleagues 

(2007), where adolescent nicotine 

enhanced self-administered 

i.v. cocaine. This difference may be 

attributable to species or the time 

between nicotine exposure and the 

start of self-administration, with 

altered dopamine dynamics during 

nicotine withdrawal promoting 

cocaine self-administration.

Table A3.1-3 Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-31

Study Species/exposure Measures Outcome/findings Comments

Mojica et al. (2014) • Sprague-Dawley rats (P28–P32 

or P86–P90)

• Rats were pretreated with 

nicotine (60 μg/kg/day, i.v.) 

for 4 days; sucrose or cocaine 

(500 μg/kg/infusion, i.v.) selfadministration for at least 

12 days, followed by extinction 

and cocaine- or sucrose-primed 

reinstatement.

• Reinforcing effects of other 

drugs of abuse

• Acquisition of cocaine selfadministration

• Nicotine pretreatment 

in adolescence enhanced 

acquisition of cocaine selfadministration, but it decreased 

reinforced/nonreinforced 

discrimination in adults.

• Adolescent exposure also 

accelerated extinction of cocaine 

seeking, with no effect on drugprimed reinstatement. 

• There was a slight enhancement 

of sucrose self-administration 

in adolescent-exposed animals, 

but no change in extinction 

or reinstatement of sucrose 

seeking.

Brief, low-dose nicotine exposure 

in early adolescence produced 

lasting enhancements of drugrelated learning. The mechanism 

may be altered dopaminergic 

signaling in the dorsal striatum in 

response to nicotine.

Pipkin et al. (2014) • Sprague-Dawley rats

• Daily injections of saline 

or nicotine (0.16 or 

0.64 mg/kg, s.c.) from P35 to 

P50 followed by a switch to 

saline or continuation of same 

nicotine dose for remainder of 

study.

• Methamphetamine 

(0.05 mg/kg, i.v.) selfadministration began ~1 week 

later.

• Reinforcing effects of other 

drugs of abuse

• Acquisition and reinstatement 

of methamphetamine selfadministration

• 0.16 mg/kg nicotine during 

the pre-exposure and onward 

produced increased responding 

for methamphetamine at FR1.

• At FR3, both groups receiving 

0.16 mg/kg nicotine (preexposure only or throughout) 

had higher methamphetamine 

intake than saline- or high-dose 

nicotine-treated animals.

• Reinstatement to 

methamphetamine seeking 

after 14 days of extinction was 

not affected by prior nicotine 

exposure.

Nicotine exposure during 

adolescence led to greater drug 

intake in adulthood, but no 

change in methamphetamine 

seeking. The mechanism for 

enhancement may be attributable 

to an additive effect of nicotine 

and methamphetamine on 

mesolimbic dopamine activity.

Table A3.1-3 Continued
A Report of the Surgeon General

A3.1-32 Appendix 3.1

Study Species/exposure Measures Outcome/findings Comments

Ehlinger et al. (2016) • Sprague-Dawley rats

• Saline or nicotine (0.05 mg/kg, 

s.c.) was co-administered with 

vehicle or the D1R antagonist 

SCH-23390 (0.05 mg/kg, 

s.c. 20 minutes before nicotine) 

every other day from P28 to 

P42.

• Tissue was collected for GolgiCox staining 1 or 21 days after 

treatment.

• Synaptic remodeling

• Nucleus accumbens MSNs

• Nicotine induced significant 

and persistent remodeling 

of dendrites on nucleus 

accumbens MSNs (i.e., increases 

in dendritic length and 

bifurcations suggesting new 

branch formation) that was 

blocked by a dopamine D1R 

antagonist.

• Nicotine also produced a 

transient, D1R-independent 

increase in spine density.

Maturation of nucleus accumbens 

MSN dendrites can be altered by 

exposure to nicotine, and druginduced remodeling was persistent 

at least into early adulthood (P63). 

The mechanism may be enhanced 

activation of the cAMP-PKA 

pathway downstream of D1Rs. No 

direct age comparison was done. 

Note: α4β2 = alpha 4 beta 2-nicotinic acetylcholine receptor subtype; α7 = alpha 7 nicotinic acetylcholine receptor subtype; α6 = alpha 6 nicotinic acetylcholine receptor 

subtype; arc = activity-regulated cytoskeleton-associated protein; [125I]α-Btx binding = measurement of binding at a7 nAChRs using the antagonist alpha-bungarotoxin;

[125I]αCtxMII binding = measurement of binding at a6 nAChRs using the antagonist alpha-conotoxin MII; [125I]A-85380 binding = measurement of binding at a4b2 nAChRs 

using the agonist A-85380; 5AP tLTP protocol = 5 action potential timing-dependent long-term potentiation protocol; 5-CSRTT = 5-choice serial reaction time task; 

BLA = basolateral amygdala; BNST = bed nucleus of the stria terminalis; cFos = protooncogene and immediate early gene used as a marker of neuronal activity; 

cAMPPKA = cyclic AMP-protein kinase A, signaling cascade; CPP = conditioned place preference; CREB = cAMP response element-binding protein; 

CRF = corticotropin-releasing factor; CTA = conditioned taste aversion; d = days; D1R = dopamine D1 receptor; FR = fixed ratio; FST = forced swim test; 

g = grams; ICR mice = Institute for Cancer Research strain of mice; ICSS = intracranial self-stimulation; i.p. = intraperitoneal; i.v. = intravenous; kg = kilogram; 

mg = milligram; mGluR2 = metabotropic glutamate receptor 2; mL = milliliter; μg = microgram; mRNA = messenger ribonucleic acid; MSN = medium spiny 

neuron; nAChR = nicotinic acetylcholine receptor; NAcshell = nucleus accumbens shell; NGFI-B = nerve growth factor-induced gene-B; PFC = prefrontal cortex; 

P85 (and similar) = postnatal day number; PVN = paraventricular nucleus of the thalamus; 86Rb+ efflux assay = measure of nicotinic acetylcholine receptor function via 

rubidium-86 ion efflux; VTA = ventral tegmental area; s.c. = subcutaneous; SMC = serial multiple choice.

Table A3.1-3 Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-33

Table A3.1-4 Human studies on the effects of nicotine exposure on fetal brain development

A. Sudden infant death syndrome and potentially related risk factors

Study Population Outcomes examined Findings

Richardson et al. 

(2009)

• 25 full-term, healthy infants: 

12 born to women who smoked 

3–20 cigarettes/day and 13 born to 

women who did not smoke

The study compared measures of arousal in 

infants of smokers and nonsmokers. Infants 

were assessed using daytime polysomnography 

at 2–4 weeks, 2–3 months, and 5–6 months 

of age. Arousal was induced by pulsatile airjet stimulation to the nostrils (trigeminal 

stimulation) during active and quiet sleep in 

supine and prone positions. Physiological and 

electroencephalogram changes were assessed, 

and arousal experiences were classified as 

subcortical activation or cortical arousal. Chisquare tests were used to conduct statistical 

comparisons. 

In tobacco-exposed infants, total arousability and 

progression from subcortical activation to cortical 

arousal were depressed at 2–4 weeks and 5–6 months 

of age. A dose-dependent relationship between cortical 

activation proportions and urinary cotinine levels was 

found for supine and prone positions at 2–3 months of 

age. Sleep position was strongly associated with arousal 

process, regardless of tobacco exposure (infants exhibited 

higher cortical arousal proportions when prone than 

when supine).

Gunnerbeck et 

al. (2011)

• 609,551 live births to Nordic mothers 

recorded in the Swedish Medical Birth 

Register

• 7,599 maternal snus users and 

58,319 smokers

Neonatal outcomes were determined from 

hospitalization ICD-10 codes P28.2 (cyanotic 

attacks of newborn); P28.3 (primary sleep 

apnea of newborn); and P28.4 (other apnea 

of newborn). Tobacco exposure data were 

collected at first antenatal visit. Associations 

between tobacco exposure and newborn apnea 

were examined and adjusted for maternal 

age, height, parity, education, infant gender, 

gestational age, being small for gestational age, 

and method of child delivery. Dual users of 

both cigarettes and snus were not included. 

Maternal snus use was significantly associated with 

infant apnea. Maternal smoking was not significantly 

associated with infant apnea after adjusting for 

gestational age, being small for gestational age, method 

of child delivery, and other variables.

Spindel & 

McEvoy (2016)

• Literature Review Examination of pulmonary effects of (a) in 

utero nicotine exposure and (b) in utero 

tobacco product exposure, and subsequent 

comparison of the health impacts of these 

exposures.

Nicotine appears to mediate most of the pulmonary 

effects of maternal smoking during pregnancy as evident 

by the similarity of effects and underlying mechanisms 

of action. The effects of maternal smoking on lung 

development are mediated by nicotinic receptors, 

changes in airway geometry, effects on airway epithelial 

cell proliferation, and oxidative mechanisms. Similarly, 

animal models show that the effects of prenatal nicotine 

exposure are also mediated by these same mechanisms. 

Therefore, both descriptive and mechanistic data support 

the likelihood that e-cigarettes affect lung development.
A Report of the Surgeon General

A3.1-34 Appendix 3.1

B. Altered corpus callosum

Study Design/population Outcomes examined Findings

Jacobsen et al. 

(2007a)

• 33 adolescents with prenatal 

exposure to tobacco (25 were 

current smokers at the time 

of the study and 8 were never 

smokers); 34 adolescents 

with no prenatal exposure 

to tobacco (14 were current 

smokers at the time of the 

study and 20 were never 

smokers)

• Participants did not have 

any other substance abuse or 

psychiatric disorders.

This study analyzed nonsmokers and smokers who were 

abstinent for 1 week before the assessment. Outcomes 

included general intelligence, reading achievement, scores 

on word recognition tasks, motor speed, depression, 

and inattention. Magnetic resonance imaging and 

fractional anisotropy were used to examine white matter 

microstructure. Prenatal tobacco exposure was ascertained 

through parental interviews. 

Smokers with prenatal exposure to tobacco 

reported more symptoms of nicotine dependence 

and depression than smokers without such 

prenatal exposure. Prenatal exposure to tobacco 

was associated with disruption of white matter 

microstructure, especially in the anterior and 

cortical regions of the internal capsule. Findings 

were significant after controlling for potential 

confounders: baseline symptoms of depression, 

gender, alcohol use, cannabis use, and prenatal 

exposure to alcohol use. 

Paus et al. (2008) • Case-control study

• 300 adolescents, 12–18 years 

of age: 146 exposed to 

maternal cigarette smoking 

during pregnancy and 

154 matched, unexposed 

controls

• Subjects were matched 

on school and maternal 

education.

• Subjects were excluded if 

they were exposed to alcohol 

during pregnancy. 

This study examined the possible effects of maternal 

smoking during pregnancy on the size and structural 

properties of the corpus callosum. The size of the corpus 

callosum and magnetization-transfer ratios were assessed 

using magnetic resonance imaging in exposed and 

unexposed participants. Prenatal exposure to tobacco 

was ascertained retrospectively through interviews with 

mothers. T-tests and analysis of variance were used to 

assess associations between exposure to tobacco and size 

of the corpus callosum. Multiple regression analysis was 

used to adjust for household income and breastfeeding.

In females, the corpus callosum, especially the 

posterior part, was smaller in those who were 

exposed to tobacco prenatally than in those who 

were not exposed. No effects were seen in males. 

Prenatal exposure to tobacco (or not) did not result 

in any differences in the myelination index for either 

gender. 

Table A3.1-4 Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-35

C. Auditory processing defects

Study Design/population Outcomes examined Findings

Kristjansson et 

al. (1989)

• 79 children, 4–7 years of age: 

52 children of women who 

smoked during pregnancy and 

27 children of women who 

were nonsmokers

• Sample was selected from 

a larger cohort study in 

Ottawa (Ontario, Canada) 

of 700 women who were 

interviewed during pregnancy 

and whose offspring were given 

neurologic and behavioral tests 

at regular intervals.

Children were given a visual vigilance task and an 

auditory vigilance task. Activity levels during testing 

were monitored. Testers were blind to exposure status. 

Information on maternal use of tobacco was collected 

during pregnancy. Associations between maternal use of 

tobacco and children’s activity scores and auditory and 

visual omission errors were analyzed using hierarchical 

linear regression. Data on maternal use of marijuana 

and alcohol, education, family income, and exposure 

to tobacco during childhood were included in adjusted 

analyses. 

Prenatal smoking was associated with increased 

activity scores (scores increased as maternal smoking 

increased). Prenatal smoking was not associated with 

auditory or visual omission errors or visual commission 

but was associated with auditory commission. 

McCartney et 

al. (1994)

• Longitudinal drug study (the 

Ottawa Prenatal Prospective 

Study)

• 110 offspring, 6–11 years 

of age 

This study assessed the potential effects of prenatal 

exposure to maternal tobacco use on central auditory 

processing. SCAN, a central auditory processing 

task, was administered blindly by testers to obtain 

information about parental tobacco use. Maternal 

cigarette use during pregnancy was assessed during 

interviews conducted throughout pregnancy. Exposure 

to secondhand smoke during pregnancy and yearly 

thereafter was assessed using a questionnaire; exposure 

to secondhand smoke during pregnancy was also 

assessed from urine cotinine. Multivariate analysis was 

conducted, and results were adjusted for family income; 

parental education; maternal age; parity; weight; 

gestational weight gain; caffeine, marijuana, and alcohol 

use; protein and calorie intake during pregnancy; child’s 

birth weight, gender, and home environment; and 

family history of hearing problems.

Maternal smoking during pregnancy was linearly 

associated with poorer performance on a central 

auditory processing task, particularly the Competing 

Words subtest. Results remained significant after 

adjusting for other drug use, demographic variables, 

and exposure to secondhand smoke during pregnancy 

and postnatally. 

Table A3.1-4 Continued
A Report of the Surgeon General

A3.1-36 Appendix 3.1

Study Design/population Outcomes examined Findings

Franco et al. 

(1999)

• 26 newborns from Belgium: 

13 were born to women who 

smoked more than nine 

cigarettes per day during 

pregnancy, and 13 were born 

to women who did not smoke 

during pregnancy.

• 42 infants, approximately 

12 weeks of age: 21 infants 

of women who smoked, and 

21 infants of women who did 

not smoke. 

White noise of increasing intensity was administered 

during rapid-eye-movement sleep. Levels of 

arousal and awakening thresholds during 9-hour, 

night-monitoring sessions were evaluated using a 

scalp electroencephalogram, electro-oculogram, 

electrocardiogram, thoracic and abdominal respiratory 

movements, and airflow by thermistors. An actigram 

recorded gross body movements. Oxygen saturation 

was recorded using a transcutaneous sensor. Outcomes 

in newborns and infants exposed and not exposed to 

tobacco were compared using Wilcoxon nonmatched 

pairs, signed-rank test, and χ2 test.

More intense auditory stimuli were needed in newborns 

and infants of smokers than in newborns and infants 

of nonsmokers. Behavioral awakening occurred less 

frequently in newborns of smokers than in those of 

nonsmokers. Results were not explained by differences 

in exposure to maternal alcohol, drug, and sedative use; 

previous sleep deprivation; body position during sleep; 

environmental light; temperature; or background noise.

Leech et al. 

(1999)

• Longitudinal study

• 608 mother/child pairs

• Children, 6 years of age, who 

were participating in the 

Maternal Health Practices and 

Child Development Project

• Mothers of lower 

socioeconomic status were 

recruited from a prenatal 

clinic; more than 50% were 

African American.

This study of 6-year-olds assessed the relationships 

between (a) prenatal exposure to alcohol, marijuana, 

tobacco, and cocaine and (b) attention and impulsivity, 

as measured by a continuous performance task. 

Child demographic factors were also included in the 

assessment.

Prenatal exposure to tobacco, marijuana, and cocaine 

was predictive of inattention and impulsivity in the 

continuous performance task. Tobacco use during the 

second and third trimesters and cocaine use during 

the first trimester predicted increased omission errors. 

Marijuana use during the second trimester predicted 

increased commission errors and fewer omission 

errors. Lower omission errors were also associated with 

younger child age, maternal work or school status, 

higher maternal hostility scores, and lower scores on 

the Stanford-Binet Intelligence Scale. 

Cowperthwaite 

et al. (2007)

• 38 fetuses of mothers who 

smoked, and 20 fetuses of 

mothers who did not smoke

• Mothers were from antenatal 

clinics in southern Canada.

Fetuses were assessed for 1 hour or more after mothers 

smoked. Assessments included spontaneous fetal heart 

rate and body movements and were conducted with 

and without maternal speaking. Outcomes in tobaccoexposed and unexposed fetuses were compared.

Spontaneous behaviors did not differ between tobaccoexposed and unexposed fetuses. Fetuses less than 

37 weeks of gestation in mothers who smoked had a 

delayed onset of response to maternal voice compared 

with fetuses in mothers who did not smoke.

Table A3.1-4 C Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-37

Study Design/population Outcomes examined Findings

Jacobsen et al. 

(2007a)

• 181 adolescent smokers and 

nonsmokers with and without 

prenatal exposure to maternal 

smoking, 13–18 years of age, 

were recruited from a local 

community.

This study assessed three kinds of attention: auditory, 

visual selective, and visual divided. A subset of 

63 subjects also underwent functional magnetic 

resonance imaging while performing auditory, visual 

selective, and visual divided attention tasks. Outcomes 

in adolescents exposed and not exposed to tobacco were 

compared. Regression models were used to adjust for 

potential confounders.

Prenatal exposure to tobacco smoke was associated with 

depressed mood at the time of the assessment, decreased 

accuracy, and slower lexical discrimination during 

auditory attention relative to visual selective and visual 

divided-attention conditions. Compared with females, 

males with prenatal exposure had a greater reduction 

in performance accuracy during auditory conditions 

relative to visual conditions. Results from magnetic 

resonance imaging indicated that subjects with prenatal 

exposure had greater activation of the right superior 

temporal gyrus during the auditory conditions and 

increased activation of the left and right lingual gyrus 

relative to subjects with no exposure to tobacco. 

Prenatal exposure to tobacco smoke was associated 

with visual attention performance accuracy. Results did 

not change after controlling for prenatal exposure to 

alcohol, cannabis, and cocaine. 

Female nonsmokers with no prenatal exposure to 

maternal smoking had the most accurate performance 

in both auditory and visual modalities; female smokers 

with prenatal exposure to tobacco performed the least 

accurately; those with prenatal or adolescent exposure 

performed intermediately. 

Key et al. 

(2007)

• 16 healthy newborn infants: 

8 born to mothers who smoked 

and 8 born to mothers who did 

not smoke

• Infants were matched on 

gender, gestational age, and 

birth weight at 48 or less hours 

old.

Event-related potentials in response to speech stimuli 

were measured to assess speech-processing ability. 

Outcomes in infants exposed and not exposed to tobacco 

were compared.

Infants of smokers demonstrated abnormal changes 

in hemispheric symmetry for speech processing. 

Compared with infants of nonsmokers, infants of 

smokers discriminated among fewer syllables, began 

the discrimination process 150 milliseconds later, and 

differentiated among fewer stimuli.

Table A3.1-4 C Continued
A Report of the Surgeon General

A3.1-38 Appendix 3.1

Study Design/population Outcomes examined Findings

Korres et al. 

(2007)

• 200 newborns undergoing 

screening for hearing 

impairment: 100 were born 

to women who smoked and 

100 to women who did not 

smoke.

• All newborns were at low risk 

for hearing loss.

TEOAEs were measured to assess hearing loss. Mothers 

were interviewed about cigarette use during each 

trimester of pregnancy; only nonsmokers and “habitual” 

smokers were enrolled. Cigarette use was categorized in 

increments of five, starting with less than five cigarettes/

day. Outcomes in newborns exposed and not exposed 

to tobacco were compared using the t-test for unpaired 

data.

The mean response and mean amplitude at 4,000 Hz of 

TEOAEs were significantly lower in newborns exposed 

to tobacco smoke than in those who were not exposed, 

supporting the conclusion that exposure to tobacco 

in utero had a modest but detrimental effect on the 

function of outer hair cells. There was no dose–response 

relationship, and all exposure groups appeared to be 

equally affected.

Kable et al. 

(2009)

• 172 mother–infant pairs 

were recruited after delivery 

from two hospitals in Atlanta, 

Georgia. 

Mother–infant pairs were recruited at delivery and 

followed longitudinally. Auditory brainstem-evoked 

responses were used to assess differences in sensory 

processing of auditory stimuli at 6 months of age. 

Planned polynomial contrasts were conducted to assess 

dose–response relationships for linear, quadratic, and 

cubic relationships. Findings were on the basis of 

tobacco use by mothers during pregnancy: smoked less 

than a half pack of cigarettes per day, a half pack to a full 

pack per day, or more than one pack per day.

Maternal smoking level was negatively associated with 

response latency, even after controlling for confounders: 

participant age, perinatal complications, and maternal 

alcohol use. 

Peck et al. 

(2010)

• 40 mother–newborn pairs 

were drawn from a cohort 

of 110 pregnant women 

from a predominantly 

lower socioeconomic-status 

population receiving prenatal 

care in Oklahoma City, 

Oklahoma.

Auditory brainstem-evoked responses were recorded 

in neonates to assess neuroelectrical activity of the 

auditory nerve following a sound stimulus. Maternal 

smoking status was determined at the first prenatal 

visit by self-reports and urine cotinine levels. Cox 

proportional hazards models were used to develop 

hazard ratios to compare rates of response in infants 

exposed and not exposed to tobacco. Several potential 

confounders were examined. 

Newborns of mothers with the highest cotinine 

concentrations responded at a rate four times that of 

nonsmoking mothers. Findings were not affected by 

postmenstrual age at testing, maternal age, maternal 

race, infant gender, education, household income, birth 

weight, or creatinine concentration. Associations with 

more moderate cotinine levels were not observed. 

Durante et al. 

(2011) 

• Cross-sectional study of 

neonates in São Paulo, Brazil

• Study group: 98 neonates 

born to mothers who smoked 

during pregnancy

• Control group: 320 neonates 

born to mothers who did not 

smoke

• Sample size = 418

TEOAEs were studied to determine the effect of 

maternal smoking during pregnancy on cochlear 

functioning. Neonates were placed in a common crib. 

A probe with a soft rubber tip was inserted into the 

external auditory meatus to record TEOAEs.

Maternal smoking during pregnancy had a negative 

effect on cochlear function, as determined by 

otoacoustic emissions testing.

Table A3.1-4 C Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-39

Study Design/population Outcomes examined Findings

Durante et al. 

(2013) 

• Cotinine levels were measured 

in youth (8–10 years of age).

• Sample size = 145

This study analyzed the effect of tobacco smoke exposure 

during childhood on cochlear physiology by measuring 

the TEOAE response levels. Comparisons were made of 

TEOAEs among youth exposed to secondhand smoke and 

those not exposed. 

Youth exposed to secondhand smoke showed lower 

response levels to auditory testing, mainly on frequencies 

of 2.8 kHz on the right and left ears and 2.0 kHz on the 

left ear, and lower signal noise response levels, mainly 

on the 1.0 kHz and 1.4 kHz frequencies, when compared 

with controls that were not exposed to secondhand 

smoke. 

Katbamna et al. 

(2013)

• 14 mother–infant pairs: 

7 smokers and 7 nonsmokers

• Infants were full term, and 

mothers were mainly from 

urban areas, with low- to 

middle-income levels and 

uncomplicated pregnancies. 

This study sought to determine relationships between 

prenatal smoke exposure-induced changes in hearing 

responses and changes in placental gene expression. 

Cochlear echo response amplitudes and auditory 

brainstem response wave latencies were examined. 

Gene microarray was used to identify upregulated and 

downregulated placental gene expression. Functional 

gene clusters with the highest enrichment scores were 

identified. 

Cochlear echo response amplitudes and auditory 

brainstem response wave latencies were reduced in 

newborns of mothers who smoked. Changes in newborn 

cochlear and auditory brainstem functions were 

correlated with changes in placental gene expression of 

a cluster of genes that modulate cochlear hair motility. 

The role of these genes in the placenta was associated 

with active contractile function in the anchoring villi 

during maternal–fetal perfusion matching. The closure 

and relaxation of the cluster of genes in placental 

anchoring villi contraction may play a similar role in 

the cochlea during sensory neurotransduction.

Kisilevsky et al. 

(2014)

• Longitudinal study of auditory 

information processing in 

fetuses

• Sample size = 167

This study measured auditory information processing in 

fetuses through 15 months of age using the Mullen Scales 

of Early Learning and MacArthur-Bates Communicative 

Development Inventory subscales. 

Findings suggest that fetal growth restriction affects 

the development of auditory system functioning and 

indicate that it may be possible to identify individual 

fetuses and newborns at risk for language deficits and 

to intervene early, when the foundation for language is 

being laid.

Table A3.1-4 C Continued
A Report of the Surgeon General

A3.1-40 Appendix 3.1

D. Appetitive behaviors: Smoking uptake, nicotine dependence

Study Design/population Outcomes examined Findings

Kandel et al. 

(1994)

• 192 adolescent girls 

followed from grades 

10 and 11 and their firstborn offspring, as selected 

from the NYS cohort

• 797 female youth from 

the NLSY cohort and their 

first-born offspring

Women who were enrolled in the NYS cohort were 

followed for 19 years, and their offspring were interviewed 

at 9–17 years of age. Women from the NLSY (a nationally 

representative sample) were enrolled as youth and 

interviewed annually; their offspring were interviewed 

at 10–18 years of age. Women and their offspring were 

interviewed about past drug use. Information about 

maternal smoking and other drug use during the 

pregnancy with the study’s offspring was ascertained 

retrospectively. Associations between prenatal tobacco 

exposure and offspring smoking and persistence of 

smoking were assessed. Logistic regression was used 

to control for maternal age, education, and smoking 

behaviors. 

The NYS analysis revealed a significant association between 

maternal smoking during pregnancy and smoking by 

their offspring 13 years later; the association was stronger 

for daughters than for sons. Controlling for alcohol and 

marijuana use during pregnancy did not change the 

findings. The association between maternal smoking 

and persistence of smoking in offspring was stronger 

for daughters than for sons. Maternal current smoking 

was examined as a potential confounder, but maternal 

smoking during pregnancy was the strongest risk factor 

for daughters’ smoking. In the NLSY analysis, maternal 

smoking during pregnancy was associated with the 

offspring’s current and persistent smoking (but not ever 

smoking), especially for daughters. 

Griesler et al. 

(1998)

• 187 mothers and their 

first-born offspring from 

the NYS cohort; mothers 

were recruited from public 

high schools in the state of 

New York.

Mothers were followed from 15–16 years of age 

to 34–35 years of age. Offspring were interviewed 

at 3–13 years of age and again at 9–17 years of 

age. Ever and past-year smoking were assessed in 

adolescent offspring. Prenatal smoking was ascertained 

retrospectively from interviews; maternal smoking in the 

first year after child delivery was also assessed. Maternal 

reports were used to assess child behavioral problems 

using scales from Schaefer and colleagues (1977) and 

Furstenberg and colleagues (1987). Outcomes in offspring 

exposed and not exposed to tobacco were compared.

Maternal smoking of at least one pack of cigarettes per day 

during pregnancy was associated with problem behaviors in 

offspring, especially in daughters. In daughters, there was 

a significant association between maternal smoking of at 

least one pack of cigarettes per day during pregnancy and 

current smoking in adolescence, even after adjusting for 

maternal current smoking, early child behavior problems, 

and maternal monitoring. In survival models, elevated 

child behavior problems were significantly associated with 

onset of smoking in girls, and maternal smoking during 

pregnancy was not significant. 

Kandel and 

Udry (1999)

• 240 mother–daughter pairs 

from the Child Health and 

Development Study were 

followed from pregnancy. 

Daughters’ smoking during adolescence and adulthood 

was assessed through interviews conducted with the 

daughters when they were 15–17 years of age and 

27–30 years of age. Maternal tobacco use during 

pregnancy was assessed by maternal self-reports and 

blood samples during pregnancy. Outcomes for daughters 

exposed and not exposed to tobacco were compared; 

results were adjusted for potential confounders.

Maternal prenatal cotinine level was not significantly 

associated with daughters’ adolescent smoking, but 

mother’s self-reported smoking was significantly associated 

with that outcome. Maternal prenatal cotinine and 

testosterone levels were positively correlated; maternal 

testosterone was significantly associated with daughters’ 

adolescent smoking. 

Table A3.1-4 Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-41

Study Design/population Outcomes examined Findings

Buka et al. 

(2003)

• 1,248 offspring from 

two samples of the 

Collaborative Perinatal 

Project, a multisite 

cohort study of more than 

50,000 women, who were 

recruited during pregnancy 

from 1959 to 1966, and 

their offspring.

• Offspring were followed 

from birth to 29 years of 

age.

Maternal smoking during pregnancy was assessed at 

each prenatal visit, offspring smoking behavior and 

nicotine dependence were obtained by interviews, per 

the Diagnostic Interview Schedule. Maternal smoking 

was categorized as none, smoked less than one pack 

of cigarettes per day, and smoked at least one pack of 

cigarettes per day. Associations between exposure and 

outcomes were assessed using logistic regression to 

generate adjusted ORs. 

Offspring of women who smoked at least one pack of 

cigarettes per day during pregnancy were more likely to 

meet DSM-III criteria for lifetime tobacco dependence than 

offspring of women who did not smoke during pregnancy. 

The odds of progressing from smoking to nicotine 

dependence were almost twice as high in offspring whose 

mothers smoked heavily during pregnancy. These findings 

remained significant after adjusting for gender and age, 

maternal socioeconomic status, and age at pregnancy. Odds 

of marijuana dependence were not significantly elevated 

among offspring of smokers.

Lieb et al. 

(2003)

• Prospective longitudinal 

cohort study

• 938 adolescents, 

14–17 years of age, 

from Munich, Germany, 

participating in the Early 

Developmental Stages of 

Psychopathology study

• Followed for 4 years

Smoking and nicotine dependence were assessed 

using the Munich Composite International Diagnostic 

Interview. Outcomes in adolescents exposed and not 

exposed to tobacco were compared. 

Children of mothers who smoked during pregnancy had 

a higher lifetime prevalence of nicotine dependence than 

children who were not exposed to tobacco during their 

mother’s pregnancy. Differences were not observed between 

sons and daughters. 

Oncken et al. 

(2004)

• 298 adult tobacco users 

seeking smoking cessation 

treatment who were able to 

report whether they were 

exposed in utero to tobacco 

This study examined the potential associations between 

maternal smoking and their offspring’s smoking 

initiation, progression of cigarette use, and current 

nicotine dependence. Participants were asked if their 

mothers smoked during pregnancy. Univariate analysis 

was used to conduct statistical comparisons.

Females exposed to tobacco in utero transitioned from 

initial to daily smoking more rapidly than females 

without in utero exposure to tobacco; the opposite was 

true for males. Measures of current use and dependence, 

including Fagerström Test for Nicotine Dependence, prior 

withdrawal, and number of past-year quit attempts, were 

associated with in utero exposure, gender, or interactions 

of exposure and gender.

Table A3.1-4 D Continued
A Report of the Surgeon General

A3.1-42 Appendix 3.1

Study Design/population Outcomes examined Findings

Cornelius et al. 

(2005)

• Prospective birth cohort 

study

• 567 mothers and their 

offspring were randomly 

selected from an urban 

hospital-based prenatal 

clinic in Pittsburgh, 

Pennsylvania.

• Followed to 14 years of age 

This study examined smoking by offspring at 14 years 

of age, amount smoked, and age at onset of smoking. 

Maternal tobacco use during pregnancy was assessed 

during each trimester of pregnancy. Smoking 

among offspring was ascertained from interviews, 

using questions adapted from the Health Behavior 

Questionnaire, and from cotinine concentration in urine. 

Associations between maternal smoking and offspring’s 

ever smoking were assessed with logistic regression. Cox 

proportional hazards was used to identify risk factors for 

age at onset of smoking.

Prenatal exposure to tobacco in the third trimester 

of pregnancy significantly predicted ever smoking in 

offspring, amount smoked, and age at onset of smoking. 

Results remained significant after adjusting for maternal 

and child psychological characteristics. Results were not 

significant, however, after adjusting for maternal current 

smoking and peer smoking. 

Roberts et al. 

(2005)

• Data from the 1958 

National Child 

Development Study and the 

1970 British Birth Survey 

This study examined associations between maternal 

smoking during pregnancy and smoking among offspring 

in adolescence and adulthood. Smoking among offspring 

at 16 and 30 or 33 years of age was assessed through 

questionnaires. Maternal smoking during pregnancy was 

assessed from a questionnaire administered at the end 

of pregnancy; smoking was defined as smoking one or 

more cigarettes per day. Multivariate logistic regression 

was used to control for potential confounders, including 

gender of offspring, birth cohort, maternal age at birth of 

offspring, and maternal and paternal smoking at 16-year 

follow-up. 

Multigroup analyses revealed that among offspring 

whose mothers smoked during pregnancy, females were 

15% more likely than males to smoke cigarettes by 16 years 

of age. Smoking at 16 years of age in female offspring 

of mothers who smoked during pregnancy doubled the 

likelihood of smoking at the 30- and 33-year follow-ups. 

Al Mamun et 

al. (2006)

• Longitudinal cohort study

• 3,058 mothers and 

offspring enrolled in 

the Mater-University of 

Queensland Study of 

Pregnancy in Brisbane, 

Australia

• Offspring followed to 

21 years of age

Mothers were asked to record their tobacco use when 

they were at 18 weeks’ gestation; 3–5 days after child 

delivery; and at the 6-month, 5-year, and 14-year followups. Smoking behaviors of offspring were compared 

on the basis of their mother’s tobacco use during 

pregnancy (never smoked, smoked before or after but not 

during pregnancy, or smoked during pregnancy) using 

regression models and controlling for confounders.

Smoking by offspring as young adults (early- or late-onset 

smoking) was significantly associated with maternal 

smoking during pregnancy compared with no maternal 

smoking, but not with maternal smoking during other 

times. Results remained significant after adjusting for 

potential confounders, including maternal age at birth of 

offspring, gender of offspring, family income, and maternal 

education.

Table A3.1-4 D Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-43

Study Design/population Outcomes examined Findings

Munafò et al. 

(2006)

• Offspring of women 

enrolled in the National 

Child Development Study, 

a cohort of the British 

population

• Followed from birth to 

adulthood

This study examined associations between maternal 

smoking during late pregnancy and the likelihood 

of smoking among offspring in adolescence and 

adulthood. Smoking among offspring was assessed from 

questionnaires administered at 16, 23, and 33 years 

of age. Maternal smoking after the fourth month of 

pregnancy was assessed from maternal self-reports at the 

end of pregnancy. The zero-inflated negative-binomial 

model was used to analyze daily cigarette counts; 

the count proportion was fit with negative binomial 

regression models, and smoking status was fit with a 

logistic regression model. Results were controlled for 

paternal socioeconomic status, maternal smoking before 

pregnancy, and parental smoking status at the 16-year 

follow-up. 

Maternal smoking in late pregnancy was associated with 

increased likelihood of male offspring being nonsmokers 

at the 16-, 23-, and 33-year follow-ups. There was no 

association observed for female offspring. 

Kandel et al. 

(2007)

• Longitudinal study

• 353 students in grades 

6–10 in public schools in 

Chicago

• One parent (usually the 

mother)

• Followed for 2 years 

Household interviews were conducted three times during 

the study period. Time to nicotine dependence criteria, 

per DSM-IV, and full dependence syndrome were assessed. 

Maternal smoking during pregnancy was assessed 

through parental interviews. Other potential risk factors 

were also assessed, including race, gender, one or both 

parents ever being nicotine-dependent, positive diagnostic 

screen for conduct disorder, pleasant experience with 

first use of tobacco, marijuana use before tobacco use, 

alcohol use before tobacco use, peer smoking, depressive 

symptoms, positive screen for anxiety disorder, academic 

performance, and pubertal stage.

Maternal smoking during pregnancy was not associated 

with time to nicotine dependence criteria, per DSM-IV, or 

time to full dependence syndrome. 

Table A3.1-4 D Continued
A Report of the Surgeon General

A3.1-44 Appendix 3.1

Study Design/population Outcomes examined Findings

Weiss et al. 

(2008) 

• Cohort study of long-term 

smokers

• Sample size = 2,827

• Long-term smokers in Utah 

and Wisconsin and by the 

NHLBI Lung Health Study

This study compared nicotine addiction, assessed via 

Fagerström Test, using a candidate-gene approach with 

neuronal nAChR subunit genes. The panel included 

common coding variants and haplotypes detected in 

eight α and three β nAChR subunit genes in European 

Americans.

Susceptibility and protective haplotypes at the 

CHRNA5-A3-B4 locus were associated with nicotine 

dependence severity (p = 2.0×10−5; odds ratio = 1.82; 

95% CI, 1.39–2.39) in subjects who began daily smoking at 

or before 16 years of age. A substantial shift in susceptibility 

versus protective diplotype frequency (AA vs. BC = 17%, AA 

vs. CC = 27%) was observed in smokers who began using by 

age 16. This genetic effect was not observed in subjects who 

began daily nicotine use after the age of 16. These results 

establish a strong mechanistic link among early nicotine 

exposure, common CHRNA5-A3-B4 haplotypes, and adult 

nicotine addiction in three independent populations of 

European origin.

O’Callaghan et 

al. (2009)

• 2,571 mothers and 

offspring enrolled in 

the Mater-University of 

Queensland Study of 

Pregnancy in Brisbane, 

Australia

The potential effects of maternal smoking during 

pregnancy on offspring nicotine dependence were 

examined using data from a longitudinal cohort whose 

offspring were followed to 21 years of age. Offspring 

completed the Composite International Diagnostic 

Interview (Auto) to assess nicotine dependence and 

withdrawal according to DSM-IV criteria. Tobacco use by 

mothers during pregnancy was assessed during prenatal 

care (never smoked, smoked before or after but not 

during pregnancy, or smoked during pregnancy). Logistic 

regression was used to control for potential confounders.

Smoking during pregnancy was significantly associated 

with offspring nicotine dependence and withdrawal 

at 21 years of age in a comparison with offspring of 

women who never smoked. This finding remained after 

adjusting for a variety of potential confounders, including 

maternal age at birth of offspring, education, duration of 

breastfeeding, parental communication, and internalizing 

or externalizing behavior problems. 

Table A3.1-4 D Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-45

Study Design/population Outcomes examined Findings

Tehranifar et 

al. (2009)

• 262 offspring of women 

enrolled in New York City’s 

National Collaborative 

Perinatal Project, a 

multisite cohort study of 

more than 50,000 women 

who were recruited during 

pregnancy from 1959 to 

1966, and their offspring

• Offspring were followed 

prospectively until 7 years 

of age, then enrolled in an 

adult follow-up study at 

approximately 41 years of 

age.

This study examined the relationship between exposure 

to secondhand smoke over the life course and adult 

smoking status. Offspring smoking status, number of 

years of smoking, age at smoking initiation, average 

daily consumption of tobacco, and age at cessation were 

assessed at adult follow-up. Maternal smoking during 

pregnancy was assessed during enrollment of mothers 

during pregnancy. Exposure to secondhand smoke in 

childhood and adulthood was assessed by self-reports of 

smokers in the home.

Current and former smokers were more likely than never 

smokers to have been exposed to prenatal tobacco and 

adult household tobacco. Results remained significant after 

adjusting for socioeconomic status.

Agrawal et al. 

(2010) 

• 1,741 offspring of 

fathers who were twins 

participating in the 

U.S. Vietnam Era Twin 

Registry and their female 

spouses 

Offspring of study participants were interviewed at 

12 and 34 years of age and were assessed for remediation, 

conduct problems, ADHD, low scholastic achievement, 

and regular smoking. Prenatal tobacco exposure was 

assessed retrospectively by interview. Associations 

between maternal smoking during pregnancy and 

outcomes were assessed using regression analysis and 

Kaplan-Meier curves for survivor function. Participants 

were tested for initiation of smoking and regular 

smoking. Potential confounders were examined, 

including ethnicity and education of parents and parental 

drug and alcohol dependence. 

Maternal smoking at any time during pregnancy was 

associated with low scholastic achievement, earlier age 

of initiation and onset of regular smoking, and ADHD in 

offspring. The association with ADHD in offspring was 

attributable to confounders (maternal ADHD, maternal 

conduct disorder, and low paternal education). 

Sullivan et al. 

(2011)

• Systematic review

• 57 studies of mothers’ 

prenatal smoking status 

in offspring using sex- and 

gender-based analyses were 

included.

This study examined whether maternal smoking during 

pregnancy influences smoking among female offspring 

more so than among male offspring. Ten studies used sexbased analyses and 51 used gender-based analyses; 4 used 

both and were counted in both groups.

Mother’s prenatal and postnatal smoking status influenced 

smoking among female offspring more so than among male 

offspring. Four of 10 studies found evidence of femalespecific effects of maternal prenatal smoking; 1 study found 

evidence of male-specific effects; 2 studies found evidence 

of effects on both males and females; and 3 studies found 

no association for either gender. 

Table A3.1-4 D Continued
A Report of the Surgeon General

A3.1-46 Appendix 3.1

Study Design/population Outcomes examined Findings

D’Onofrio et 

al. (2012)

• Two separate samples 

from the United States and 

Sweden

• Sample 1: A representative 

sample of 6,094 adolescent 

offspring of the women in 

the NLSY (1979)

• Sample 2: All (1,187,360) 

offspring born in Sweden 

from 1983 to 1995

Outcomes included self-reports from adolescents about 

their use of cigarettes and other substances, age at onset 

of cigarette and substance use before or after 14 years of 

age, and convictions and hospitalizations for alcohol—or 

drug-related problems. Collection of maternal tobacco 

use, covariates, and outcomes among offspring varied 

by study. Cox proportional hazards survival analysis was 

used for all outcomes, which were right-censored; logistic 

regression models were used for dichotomous outcomes. 

Maternal smoking during pregnancy was associated with 

adolescent alcohol use, cigarette use, marijuana use, 

substance-related driving convictions, narcotic convictions, 

and drug-related hospitalizations. In an analysis of 

differentially exposed siblings using data from Sweden, the 

associations were attenuated and no longer statistically 

significant. 

Rydell et al. 

(2012)

• 3,020 Swedish youth, 

11–18 years of age, in the 

Children’s Smoking and 

Environment in Stockholm 

County Study (the BROMS)

This study examined potential associations between 

maternal smoking during pregnancy and adolescent 

tobacco uptake, nicotine dependence, and withdrawal 

symptoms. Adolescent outcomes were assessed through 

self-administered questionnaires of the youth. Parental 

questionnaires were used to retrospectively assess 

prenatal exposure to tobacco. Cox regression was 

employed to analyze the onset of tobacco use. Logistic 

regression was used to study cravings, withdrawal 

symptoms, and tobacco consumption at 17 years of 

age. Results were adjusted for potential confounders, 

including parental education and postnatal tobacco use. 

Girls with prenatal exposure to tobacco had two to 

three times the odds of experiencing a high number of 

withdrawal symptoms, craving for tobacco, and heavy 

tobacco use (five or more cigarettes smoked or snus dips 

per day), but not onset of use. No significant associations 

were observed among boys. 

Weden and 

Miles (2012)

• 6,349 participants, 

14–25 years of age, from 

the Children and Young 

Adults of the NLSY

Intergenerational transmission of smoking was assessed. 

A panel survey of all offspring of women in a populationrepresentative cohort was used; children born to 

participating women were followed longitudinally. Youth 

smoking trajectories were constructed using mixture 

latent trajectory analysis. 

The classes of smoking trajectory were early onset, early 

experiment, late onset, and nonsmoker. Analysis of the 

three smoking trajectories indicated that in all cases, youth 

whose mothers smoked during pregnancy were more likely 

to develop the trajectory than to become nonsmokers. 

Table A3.1-4 D Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-47

Study Design/population Outcomes examined Findings

Stroud et al. 

(2014)

• 1,086 mother–offspring 

pairs enrolled in the 

Collaborative Perinatal 

Project, a multisite 

cohort study of more than 

50,000 women who were 

recruited during pregnancy 

from 1959 to 1966, and 

their offspring

This study examined the association between maternal 

smoking during pregnancy, offspring nicotine 

dependence, and the potential mediating roles of 

prenatal glucocorticoid and androgens. Lifetime nicotine 

dependence among offspring was assessed by interviews 

during 40 years of follow-up and was based on DSM-IV 

criteria. Maternal smoking during pregnancy was 

assessed during prenatal visits. Bivariate associations 

were estimated with polychoric, polyserial, and Pearson 

correlations. A causal steps approach was used to examine 

potential mediators using multivariate logistic regression 

analysis. Potential confounders were examined, including 

gravidity and parity, maternal age at child delivery, race 

and ethnicity, socioeconomic status, maternal use of 

other drugs, and maternal history of treatment for mental 

illness. 

Cotinine-validated maternal tobacco use during pregnancy 

was significantly associated with nicotine dependence in 

daughters. Results remained significant after adjusting for 

potential confounders, including maternal demographic 

and obstetrical factors, alcohol and drug use, and maternal 

mental disorders. 

Taylor et al. 

(2014)

• 6,484 offspring, 

14–16 years of age, of 

participants in the Avon 

Longitudinal Study of 

Parents and Children 

Study included comparisons of associations between 

(a) maternal and partner smoking during pregnancy 

and (b) smoking initiation among offspring, using 

partner smoking as a negative control and a Mendelian 

randomization analysis (n = 1,020) using a genetic variant 

in mothers of European ancestry only. Parental smoking 

during pregnancy was assessed during pregnancy; status 

of smoking among offspring was assessed at 14–16 years 

of age by questionnaire (nonsmoker, experimenter, lateonset regular smoker, and early-onset regular smoker). 

Multinomial logistic regression was used to assess 

associations between parental smoking and outcomes 

among offspring, and results were adjusted for potential 

confounders. 

Maternal and partner smoking were associated with 

the smoking status of offspring, even after adjusting 

for confounders, but there was no clear dose–response 

relationship with the level of parental smoking. Maternal 

rs1051730 genotype (associated with heavy smoking) was 

not associated with smoking among offspring. 

Table A3.1-4 D Continued
A Report of the Surgeon General

A3.1-48 Appendix 3.1

Study Design/population Outcomes examined Findings

Shenassa et al. 

(2015)

• 1,783 adult offspring 

of women who were 

enrolled in the Providence 

and Boston sites of the 

Collaborative Perinatal 

Project (a multisite 

cohort study of more than 

50,000 women who were 

recruited during pregnancy 

from 1959 to 1966, and 

their offspring) and also 

enrolled in the New 

England Family Study 

This study examined maternal smoking during pregnancy 

and risk of nicotine dependence among offspring. Followup studies of offspring were conducted using multistage 

sampling, which oversampled families with multiple 

siblings and siblings discordant for maternal smoking 

during pregnancy. Maternal smoking during pregnancy 

was assessed during prenatal study visits. Smoking among 

offspring was assessed by the Life Interview of Smoking 

Trajectory Questionnaire. Lifetime nicotine dependence 

was based on DSM-IV criteria and assessed using the 

Composite International Diagnostic Interview. Logistic 

regression models were used to explore independent 

associations between maternal smoking during pregnancy 

and offspring smoking, nicotine dependence, and 

marijuana use. 

Maternal smoking during pregnancy was significantly 

associated with progression from weekly smoking 

to nicotine dependence but not weekly smoking or 

progression to marijuana dependence. Within-family and 

between-family analyses supported an effect of maternal 

smoking during pregnancy on progression to nicotine 

dependence. 

E. Appetitive behaviors: Use of other substances

Study Design/population Outcomes examined Findings

Fergusson et al. 

(1998)

• Birth cohort of 

1,265 children born in 

Christchurch, New Zealand

• Followed to 18 years of age

Outcomes were assessed at 16–18 years of age 

and included conduct disorder, major depression, 

anxiety, and substance use. Outcomes were 

compared between adolescents exposed and not 

exposed to tobacco in utero.

Exposure to tobacco in utero was significantly associated 

with psychiatric symptoms for conduct disorder, depression, 

substance abuse, and alcohol abuse. The association with 

conduct disorder remained significant after adjusting for 

potential confounders, including lower socioeconomic status, 

impaired child-rearing behaviors, and parental and family 

problems. Effects were more pronounced among males than 

among females. 

Table A3.1-4 D Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-49

Study Design/population Outcomes examined Findings

Weissman et al. 

(1999)

• 50 offspring of mothers 

who smoked during 

pregnancy and 97 offspring 

of mothers who did not 

smoke during pregnancy

• Offspring were followed for 

10 years.

This study examined associations between 

maternal smoking during pregnancy and ADHD, 

conduct disorder, and substance abuse among 

offspring. Psychiatric diagnostic assessments were 

conducted by interview with parents and offspring 

using the Schedule for Affective Disorders and 

Schizophrenia—Lifetime Version. Maternal smoking 

during pregnancy was assessed retrospectively 

by interview. Proportional hazards regression 

models were used to examine associations between 

offspring’s exposure to tobacco and psychopathology. 

Results were adjusted for demographic factors, 

parental psychopathological conditions, pregnancy 

risk factors, and family parenting risk factors.

Maternal tobacco use during pregnancy was significantly 

associated with prepubertal onset of conduct disorder in 

boys, and adolescent onset of drug dependence in girls. These 

findings were not explained by maternal substance use during 

pregnancy, parental psychiatric diagnosis, family risk factors, 

prenatal and early-developmental history of offspring, postnatal 

maternal smoking, or smoking among offspring. 

Porath and 

Fried (2005)

• 152 adolescents, 

16–21 years of age, born of 

mothers who participated 

during pregnancy in 

the Ottawa Prenatal 

Prospective Cohort Study

This study examined associations between maternal 

cigarette smoking and marijuana use during 

pregnancy and risk of initiation and regular use 

of substances by offspring. Outcomes included 

initiation and daily or regular use of tobacco and 

marijuana. Maternal tobacco use was ascertained 

during pregnancy. Logistic regression was used 

to compare outcomes in exposure groups while 

controlling for potential confounders, including 

parity, gender, maternal use of alcohol and caffeine 

during pregnancy, family income, parental 

education, postnatal exposure to secondhand smoke, 

exposure to marijuana smoke, and age of offspring 

at time of testing. 

Offspring of mothers who smoked cigarettes during pregnancy 

had increased risks for initiation of cigarette smoking during 

adolescence. Findings were more pronounced for males than 

females, and they remained significant after adjusting for 

confounders.

Table A3.1-4 E Continued
A Report of the Surgeon General

A3.1-50 Appendix 3.1

Study Design/population Outcomes examined Findings

Nomura et al. 

(2011)

• 1,625 offspring of women 

who were enrolled in the 

Providence (Rhode Island) 

and Boston (Massachusetts) 

sites of the Collaborative 

Perinatal Project 

(a multisite cohort study of 

more than 50,000 women 

who were recruited during 

pregnancy from 1959 to 

1966 and their offspring) 

and also enrolled in the 

New England Family Study

• Offspring were followed 

from birth to 40 years of 

age and older.

This study examined the association between 

maternal smoking during pregnancy and lifetime 

risk of alcohol use disorder in offspring. Lifetime 

risk of alcohol use disorder was assessed using 

DSM-IV criteria. Maternal smoking status was 

assessed during prenatal visits. Survival analysis 

techniques were used to examine differences 

between proportions of subjects free from alcohol 

use disorders. Potential mediating factors were 

explored, including low birth weight, neurologic 

abnormalities, poor academic functioning, and 

behavioral dysregulation.

Offspring of mothers who smoked 20 or more cigarettes 

per day during pregnancy had greater risk of alcohol use 

disorder than offspring of mothers who did not smoke during 

pregnancy. No excess risk was noted in offspring of mothers 

who smoked fewer than 20 cigarettes per day during pregnancy. 

Maternal smoking during pregnancy was associated with 

neurobehavioral problems in infancy and childhood, which, 

in turn, were associated with increased risk for alcohol use 

disorder in adulthood. 

Huang et al. 

(2013) 

• Six- to 9-week-old male 

mice, kept in plastic cages 

on a 12-hour day/night 

cycle in groups of five with 

ad libitum food and water

This study examined whether the metaplastic effect 

of nicotine on cocaine also applies in the amygdala. 

The study examined whether exposure to nicotine 

prior to exposure to cocaine enhances LTP in the 

amygdala. Short-term (1 day) and long-term (7 days) 

exposure to nicotine prior to cocaine exposure was 

examined.

Pretreatment with nicotine (i.e., exposure to nicotine prior to 

cocaine exposure) enhanced LTP in response to cocaine in the 

amygdala in both groups (short and long). The association is 

unidirectional in that pretreatment with cocaine followed by 

exposure to nicotine does not elicit the same response in the 

amygdala. After 40 days of nicotine cessation, the LTP response 

can no longer be generated.

Table A3.1-4 E Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-51

F. Appetitive behaviors: Overweight and obesity

Study Design/population Outcomes examined Findings

Gilman et al. 

(2008)

• 52,919 offspring from the 

Collaborative Perinatal 

Project, a multisite 

cohort study of more than 

50,000 women who were 

recruited during pregnancy 

from 1959 to 1966 and their 

offspring

This study assessed maternal tobacco use during pregnancy 

and 14 developmental outcomes in offspring (e.g., obesity, 

intelligence, academic achievement, conduct problems, 

and asthma). Cognition was assessed in offspring at 4 years 

of age (Stanford-Binet Intelligence Scale) and 7 years of 

age (Wechsler Intelligence Scale for Children). Academic 

performance was assessed at 7 years of age (Wide Range 

Achievement Test). Neurologic “soft signs” were assessed 

by physician examination. Behavioral observations were 

made at 7 years of age through psychological assessments; 

conduct problems were determined on the basis of 

consistency and stability across samples. Body mass index 

was recorded at 7 years of age. Maternal smoking during 

pregnancy was assessed during prenatal visits. Generalized 

estimating equations and linear and logistic regression 

analyses were used; matched analyses among siblings were 

conducted. 

Maternal smoking was associated with overweight at 

birth and with childhood overweight in unadjusted 

and adjusted models. Associations between maternal 

smoking and 12 other outcomes that were significant 

in unadjusted analyses were no longer significant after 

adjusting for potential confounders, including race and 

ethnicity, parental ages, household crowding, parental 

history of psychiatric and substance use disorders, 

maternal employment, and socioeconomic status.

Oken et al. 

(2008)

• Systematic review and metaanalysis

• 84,563 children enrolled in 

14 observational studies

This study examined associations between maternal 

smoking during pregnancy and childhood overweight. 

Children whose mothers smoked during pregnancy 

were at greater risk for overweight at 3–33 years of age 

compared with children whose mothers did not smoke 

during pregnancy, even when adjusted ORs were used. 

Meta-analysis generated a pooled adjusted OR of 1.50 

(95% CI, 1.36–1.65).

Iliadou et al. 

(2010)

• Population-based cohort study

• 124,203 singleton males born 

to Nordic women in Sweden

• Males followed to 18 years of 

age

This study examined associations between maternal 

smoking during pregnancy and childhood overweight and 

obesity in young adult Swedish males born of mothers 

who did and did not smoke during pregnancy. Associations 

among siblings were also examined, controlling for 

common genes and shared environment. 

The risk of overweight was higher in sons of mothers 

who smoked during pregnancy than in sons of mothers 

who did not smoke during pregnancy. However, after 

stratifying for maternal smoking across two subsequent 

male pregnancies, the risk in the second pregnancy was 

higher only if the mother smoked during both male 

pregnancies, suggesting that increased risk is explained 

by familial factors.

Table A3.1-4 Continued
A Report of the Surgeon General

A3.1-52 Appendix 3.1

Study Design/population Outcomes examined Findings

Ino (2010) • Systematic review and metaanalysis

• 17 studies, ranging from 

252 to 34,866 participants

This study examined associations between maternal 

smoking during pregnancy and childhood obesity in 

offspring. 

All studies showed a positive association between 

maternal smoking during pregnancy and childhood 

obesity. Meta-analysis using data from 16 studies of 

offspring, 3–33 years of age, generated a pooled adjusted 

OR of 1.64 (95% CI, 1.42–1.90). Results were significant 

after adjusting for publication bias. Maternal obesity, 

low socioeconomic status, low birth weight, and absence 

of breastfeeding did not appear to confound results. 

Lotfipour et 

al. (2010)

• 423 adolescents from the 

Saguenay Youth Study in 

Quebec, Canada 

This study examined associations between prenatal 

exposure to maternal cigarette smoking and outcomes 

in offspring. Cigarette smoking and substance use by 

offspring were assessed by questionnaire, and the volume of 

dopamine-rich regions of the striatum was measured with 

T1-weighted magnetic resonance imaging. The study also 

assessed potential interactions between prenatal exposure 

to maternal cigarette smoking and a polymorphism in 

the offspring in the α6 nAChR gene and the outcomes of 

interest. Offspring exposed and not exposed to maternal 

cigarette smoking were matched for maternal education.

Significant interactions exist between prenatal exposure 

to maternal cigarette smoking and the α6 nAChR 

SNP in the offspring in predicting adolescent lifetime 

smoking and substance use. The α6 nAChR subunit 

could modify these effects. 

Weng et al. 

(2012)

• Systematic review and metaanalysis

• Prospective studies of children 

followed from birth to at least 

2 years of age, seven of which 

examined maternal smoking 

and childhood overweight 

This study examined risk factors, including maternal 

smoking during pregnancy, for childhood overweight 

identifiable during infancy.

Maternal smoking during pregnancy was associated with 

childhood overweight in all seven of the studies where 

this was examined. Meta-analysis using data from these 

seven studies generated a pooled adjusted OR of 1.47 

(95% CI, 1.36–1.73). 

Behl et al. 

(2013)

• 83 studies of children 1 year of 

age or older 

This review of epidemiologic studies examined maternal 

smoking during pregnancy and obesity in offspring. 

Evidence (based on human and animal research) 

supports the conclusion that maternal smoking 

increases risk of obesity or overweight in offspring, but 

contributions from unmeasured residual confounding 

cannot be ruled out. Data were insufficient to assess 

contributions of exposure to secondhand smoke after 

child delivery. 

Table A3.1-4 F Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-53

Study Design/population Outcomes examined Findings

La Merril et 

al. (2015) 

• Prospective study of 

daughters, 44–54 years of age 

• Sample size = 1,801

This study aimed to identify whether parental tobacco 

smoking during gestation was associated with risk of 

diabetes mellitus.

Prenatal maternal smoking had a stronger association 

with daughters’ diabetes mellitus risk than did prenatal 

paternal smoking. Estimates of the effect of parental 

smoking were unchanged when further adjusted by 

daughters’ birth weight or current body mass index.

Harrod et al. 

(2015) 

• Mother–offspring pairs were 

analyzed in the longitudinal 

Healthy Start study.

• Sample size = 670

This study examined associations between exposure to 

prenatal smoking and early-life changes in fat mass (FM), 

fat-free mass (FFM), and anthropometrics.

Exposure to prenatal smoking was significantly 

associated with reduced neonatal FM (p = 0.007) and 

FFM (p = 0.02). Overall, exposure to prenatal smoking 

was significantly associated with rapid postnatal growth, 

which may increase the offspring’s risk of metabolic 

diseases.

Mourtakos 

et al. (2015) 

• A random sample of 

5,125 children was extracted 

from a national database and 

matched with their mothers.

This study investigated the association between gestational 

weight gain, maternal age, and lifestyle habits (e.g., physical 

activity, smoking, alcohol consumption) during pregnancy 

and body mass index of the offspring at the age of 8.

Maternal behaviors, including smoking status during 

pregnancy, were significantly associated with obesity in 

the offspring at the age of 8.

Bao et al. 

(2016) 

• Self-reported survey

• Sample size = 15,665 singleton 

pregnancies from 

10,152 women

This study examined fetal exposure to parental smoking and 

gestational diabetes in daughters.

Maternal heavy smoking (≥25 cigarettes/day) during 

pregnancy was associated with higher risk of gestational 

diabetes in the daughter.

Table A3.1-4 F Continued
A Report of the Surgeon General

A3.1-54 Appendix 3.1

G. Appetitive behaviors: Altered size and sensitivity of brain reward systems

Study Design/population Outcomes examined Findings

Toro et al. 

(2008)

• 314 adolescents: 155 exposed 

and 159 not exposed to 

tobacco smoke in utero 

This study examined the effects of maternal 

smoking during pregnancy on several outcomes in 

offspring: thickness of the cerebral cortex; general 

intelligence (using the Wechsler Intelligence Scale 

for Children); and positive youth development. 

Maternal smoking during pregnancy was assessed 

retrospectively through a maternal interview. 

Thickness of the cerebral cortex was measured using 

magnetic resonance imaging. Analyses of cortical 

thickness were performed using mean values for 

each of 33 regions and employing thickness values 

determined at each of 163,842 vertices. Positive 

youth development was assessed using the five Cs 

model (competence, confidence, character, social 

connection, and caring or compassion). Age was 

included as a covariate in each analysis. Adolescents 

exposed and not exposed in utero to tobacco smoke 

were matched on maternal education.

Prenatal exposure to maternal smoking was associated with 

thinner orbitofrontal, middle frontal, and parahippocampal 

cortices, especially in females. Based on these findings, 

the score for “caring” was examined specifically; there was 

a significant negative correlation between the thickness 

measurements of the orbitofrontal cortex and scores for 

“caring” in females who were exposed in utero to maternal 

smoking.

Lotfipour et al. 

(2009)

• 597 adolescents 

(275 sibships), 12–18 years 

of age, from the Saguenay 

Youth Study in Quebec, 

Canada 

This study assessed the potential involvement of the 

orbitofrontal cortex in mediating the relationship 

between maternal smoking during pregnancy 

and substance use in offspring. Substance use by 

offspring was assessed through a questionnaire, and 

thickness of the orbitofrontal cortex was measured 

with T1-weighted magnetic resonance imaging. 

Behavioral data were analyzed using hierarchical 

linear modeling. Potential confounders were explored, 

including breastfeeding status and parent-based 

predictors of substance use. Cortical thickness was 

analyzed in 314 adolescents; associations between 

genotype, behavior, and cortical thickness were 

examined using linear models that included potential 

interactions. Adolescents exposed and not exposed to 

maternal smoking during pregnancy were matched 

on maternal education.

Prenatal exposure to tobacco was significantly associated 

with substance use in offspring. Results did not change after 

controlling for potential confounders, including birth weight, 

breastfeeding status, maternal alcohol use during pregnancy, 

and parental history of antisocial behavior. Among exposed 

adolescents, the likelihood of drug experimentation was 

associated with the degree of thinning in the orbitofrontal 

cortex. 

Table A3.1-4
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-55

Study Design/population Outcomes examined Findings

Haghighi et al. 

(2013)

• 378 adolescents, 13–19 years 

of age, from the Saguenay 

Youth Study in Quebec, 

Canada 

This study assessed associations between prenatal 

exposure to maternal cigarette smoking and enhanced 

fat intake and risk of obesity in offspring. Exposure 

to maternal smoking was defined as smoking more 

than one cigarette per day in the second trimester 

of pregnancy vs. no exposure at any time during the 

12 months preceding and throughout pregnancy. 

Subtle variations in brain regions involved with 

reward processing were studied with magnetic 

resonance imaging; regions included the amygdala, 

nucleus accumbens, and orbitofrontal cortex. 

Offspring exposed and not exposed to maternal 

smoking during pregnancy were matched for 

maternal education. Regression models adjusted for 

gender, age, and height. 

Prenatal exposure to maternal smoking was significantly 

associated with higher total body fat and fat intake, as well as 

lower volume of the amygdala. Amygdala volume correlated 

inversely with fat intake. 

Müller et al. 

(2013)

• A subgroup of 177 healthy 

adolescents, 13–15 years of 

age, with prenatal exposure 

to maternal tobacco use, and 

177 matched controls not 

exposed to maternal tobacco 

use were recruited for the 

IMAGEN study—a European 

multicenter study of 

impulsivity, reinforcement 

sensitivity, and emotional 

reactivity in adolescents. 

This study examined the effects of maternal tobacco 

use during pregnancy on offspring’s brain response 

during reward processing. Response to reward in 

the ventral striatum was assessed with functional 

magnetic resonance imaging. Adolescents exposed to 

prenatal maternal tobacco use and those not exposed 

were matched for gender, maternal education level, 

and imaging site. 

Prenatal exposure to maternal tobacco use was associated 

with a weaker response in the ventral striatum during 

reward anticipation among those exposed than in unexposed 

controls. There were no differences in responsivity of the 

ventral striatum to the receipt of a reward. 

Table A3.1-4 G Continued
A Report of the Surgeon General

A3.1-56 Appendix 3.1

H. Attention and cognition, behavioral disorders

Study Design/population Outcomes examined Findings

Linnet et al. 

(2003)

• Systematic review

• 24 studies of maternal 

tobacco use during 

pregnancy

• Offspring were 4–36 years 

of age.

This study examined behavioral outcomes in offspring 

related to ADHD in childhood and prenatal exposure to 

nicotine, alcohol, and caffeine, and psychosocial stress. 

Only studies using validated diagnostic or screening 

instruments for ADHD and ADHD symptoms were 

included. No meta-analysis was performed.

A greater risk of ADHD-related disorders was observed in 

children whose mothers smoked during pregnancy. Many 

studies had methodologic shortcomings, such as recall bias, 

limited assessments of exposure, low statistical power, or 

insufficient control for confounders.

Langley et al. 

(2005)

• 13 studies reviewed

• Participants were 

5–12 years of age.

This review examined 13 population-based 

epidemiologic studies of ADHD-related behaviors and 

maternal smoking during pregnancy. A pooled OR for 

case control studies (n = 5) was calculated.

The majority of studies (n = 11) identified maternal smoking 

during pregnancy as a risk factor for ADHD behaviors; the 

pooled OR for case control studies was 2.39 (95% CI, 1.61–3.52). 

Methodologic limitations included misclassification of exposure 

and an inability to control for all relevant confounders and 

genetic factors.

D’Onofrio et 

al. (2008)

• Nationally representative 

sample of 6,283 mothers, 

who were enrolled at 

14–21 years of age in the 

NLSY, and their offspring, 

who were 4–10 years of 

age 

This study assessed associations between maternal 

smoking during pregnancy and conduct problems, 

oppositional defiance problems, and attention deficit/

hyperactivity problems in offspring. Offspring were 

compared with siblings who differed in their prenatal 

exposure to tobacco. Smoking during the most recent 

pregnancy was assessed by interviews at each wave of 

the study.

There was no effect of maternal smoking during pregnancy 

on conduct problems or oppositional defiance problems in the 

offspring. There was a modest association with attention deficit/

hyperactivity problems that was attenuated by methodologic and 

statistical controls.

Bennett et al. 

(2009)

• 18 children, 12 years 

of age, 7 with prenatal 

exposure to tobacco, and 

11 not exposed prenatally 

The brain function of children exposed and not 

exposed to tobacco prenatally was assessed during a 

go/no-go response inhibition task using event-related 

functional magnetic resonance imaging. Outcomes 

for exposed and not exposed children were compared, 

and results were controlled for prenatal exposure to 

alcohol, neonatal medical problems, environmental 

risks, IQ, current exposure to secondhand smoke, and 

handedness.

Children exposed to tobacco prenatally showed greater 

activation in a relatively large and diverse set of regions of 

the brain, including left frontal, right occipital, and bilateral 

temporal and parietal regions. Unexposed children showed 

activation in the cerebellum, which is important for attention 

and motor preparation. Tobacco-exposed children showed 

inefficient recruitment of regions required for response 

inhibition.

Table A3.1-4 Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-57

Study Design/population Outcomes examined Findings

Thapar et al. 

(2009)

• 815 families of offspring 

conceived through 

assisted reproductive 

technologies and born 

to either related or 

genetically unrelated 

women

• Women were recruited 

through fertility clinics in 

the United Kingdom and 

the United States.

This study examined associations between maternal 

smoking during pregnancy and ADHD in offspring 

among related and unrelated mother–offspring pairs. 

Offspring were assessed at 4–11 years of age through 

parental questionnaires and antenatal records. Multiple 

regression analysis was used to control for maternal 

alcohol use, multiple-birth status, birth weight, 

gestational age, parental ADHD symptoms, family 

income, and age and gender of offspring.

The associations between maternal smoking during pregnancy 

and ADHD symptoms in offspring were much stronger for 

related pairs than unrelated pairs, but this was not the case for 

maternal smoking and birth weight, where associations were 

similar for related and unrelated pairs.

Boutwell and 

Beaver (2010)

• An estimated 3,300 U.S. 

children from the Early 

Childhood Longitudinal 

Study, Birth Cohort

• Children were followed 

from 9 months to 4 years 

of age. 

Behavioral, cognitive, and emotional development 

were assessed using the Preschool and Kindergarten 

Behavior Scales—Second Edition. Maternal smoking 

was assessed from the birth certificate and categorized 

as “yes” or “no.” Data on potential confounders 

included maternal antisocial behaviors, substance 

abuse (alcohol and other drugs), depression, and 

education attainment; the child’s race and gender, 

rearing environment, complications of labor and 

delivery, and Apgar scores; and paternal antisocial 

behavior, substance abuse, and depression. Propensity 

score-matching analysis was performed using 

14 covariates. 

Before matching on propensity scores, subjects differed on 

11 of 14 covariates. Prior to being placed in matched groups, 

children of mothers who smoked during pregnancy exhibited 

higher levels of externalizing problem behaviors than children of 

mothers who did not smoke during pregnancy. After placement 

in matched groups, prenatal exposure to tobacco no longer 

predicted externalizing behavior problems. 

Table A3.1-4 H Continued
A Report of the Surgeon General

A3.1-58 Appendix 3.1

Study Design/population Outcomes examined Findings

Wakschlag et 

al. (2010)

• Pregnancy cohort study

• 139 adolescent offspring 

from the Maternal Infant 

Smoking Study of East 

Boston

The study tested whether a functional polymorphism in 

the gene encoding the enzyme monoamine oxidase A 

(MAOA) interacts with prenatal exposure to tobacco 

to predict outcomes in offspring. Outcomes included 

antisocial behavior (conduct disorder symptoms and 

hostile attribution bias). Maternal tobacco exposure 

(active smoking and exposure to secondhand smoke) 

was assessed via questionnaire and serum cotinine 

levels during pregnancy. Covariates included family 

income, age of offspring, prenatal maternal alcohol 

use, parental antisocial behavior, and harsh parenting. 

Linear mixed models were used to examine conduct 

disorder symptoms, and linear models were used 

to examine hostile attribution score. Permutation 

methods were used to test for gene–prenatal tobacco 

exposure interactions. 

Boys with low-activity MAOA 5’ uVNTR genotype who were 

exposed to tobacco prenatally were at increased risk for 

conduct disorder symptoms. Girls with high-activity MAOA 

uVNTR genotype who were exposed to tobacco prenatally were 

at increased risk for conduct disorder symptoms and hostile 

attribution bias. 

Lavigne et al. 

(2011)

• Convenience sample of 

679 children, 4 years of 

age, who were recruited 

from primary care 

practices throughout 

Cook County, Illinois, and 

from public schools with 

preschool programs in 

Chicago 

Internalizing and externalizing behavioral problems 

and negative temperament were assessed using several 

modalities: DISC-YC, CSI ADHD scales, DISC-YC 

generalized anxiety scale and CSI generalized anxiety 

scale (for anxiety symptoms); and DISC-YC major 

depression scale, CSI major depression scale, and 

CSI dysthymia scale (for depressive symptoms). For 

ADHD, the inattentive (ADHD-I) and hyperactive 

(ADHD-H) subtypes were examined separately, as was 

the combined type (ADHD-C), using the Children’s 

Behavior Questionnaire negative affectivity scale. 

Mothers were interviewed about their smoking status 

during pregnancy. Outcomes in children exposed and 

not exposed to tobacco through maternal smoking 

during pregnancy were compared using analysis of 

variance and linear regression; results were adjusted 

for socioeconomic status, life stress, family conflict, 

maternal depression, maternal support/scaffolding, 

mother–child attachment, and child negative affect and 

effortful control. Smoking status during pregnancy was 

assessed through interviews with the expectant mothers. 

Smoking during pregnancy was associated with ADHD in 

an unadjusted analysis, but not after correcting for multiple 

comparisons. Smoking during pregnancy was not associated 

with ADHD-H or ADHD-C. Smoking during pregnancy was 

not associated with symptoms of oppositional defiant disorder, 

anxiety, or depression in offspring.

Table A3.1-4 H Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-59

Study Design/population Outcomes examined Findings

Wakschlag et 

al. (2011)

• Pregnancy cohort study

• 211 adolescent offspring 

(30 sibling pairs) from the 

Maternal Infant Smoking 

Study of East Boston 

This study examined potential associations between 

prenatal exposure to maternal tobacco use and 

disruptive behavior outcomes (aggression dimension, 

noncompliance dimension, temper loss dimension, 

and low concern dimension) in offspring. Exposure to 

tobacco through the mother (via active smoking and 

exposure to secondhand smoke) was assessed through 

a questionnaire and serum cotinine levels during 

pregnancy. Covariates included offspring’s exposure 

to secondhand smoke and age, parental antisocial 

behavior, and family adversity. Multilevel repeatedmeasures models were used to examine effects of 

exposure while controlling for age and gender of 

offspring, parental antisocial behavior, family adversity, 

and current exposure to secondhand smoke. 

Prenatal exposure to maternal tobacco use was significantly 

associated with noncompliance and aggression dimensions, 

even after adjusting for potential confounders. There were no 

significant associations with temper loss or low concern for 

others. Maternal responsive engagement did not moderate the 

effects of the exposure on the average estimate of disruptive 

behavior, but there was a significant interaction between 

paternal responsive engagement and prenatal tobacco exposure 

for an average estimate of disruptive behavior. 

Langley et al. 

(2012)

• 8,324 offspring from the 

Avon Longitudinal Study 

of Parents and Children in 

the United Kingdom

• Offspring were followed 

from gestation to late 

adolescence.

ADHD symptoms among offspring were assessed at 

91 months of age through parental completion of the 

Development and Wellness Being Assessment. Maternal 

smoking status was assessed at 18 and 32 weeks of 

gestation (paternal smoking at 18 weeks). Associations 

between maternal smoking during pregnancy and 

paternal smoking and ADHD in offspring were 

assessed using regression analysis to explore causal vs. 

household-level confounding. Potential confounders 

included gender of offspring, ethnicity, singleton/

twin status, maternal alcohol use during pregnancy, 

education, and parental occupation.

ADHD symptoms among offspring were associated with 

maternal smoking during pregnancy and paternal smoking. 

Paternal exposure was associated independent of maternal 

exposure, suggesting genetic or household-level confounding 

rather than a causal relationship. 

Latimer et al. 

(2012)

• 47 studies of prenatal 

exposure to maternal 

smoking present 

antenatally and in the 

first 4 years of life 

This review examined studies on ADHD; oppositional 

defiant disorder; conduct disorder; and deficits in 

attention, motor control and perception, and reactive 

attachment disorder. 

Eight high-quality, large, population-based cohort studies, in 

which potential confounding was well controlled, found a link 

between prenatal maternal smoking and increased risk of ADHD 

in offspring. 

Table A3.1-4 H Continued
A Report of the Surgeon General

A3.1-60 Appendix 3.1

Study Design/population Outcomes examined Findings

McCrory and 

Layte (2012)

• 7,505 children, 9 years of 

age, participating in the 

Growing Up in Ireland 

study 

Children’s behavioral problems were assessed using 

the Strengths and Difficulties Questionnaire across 

instruments for parents and teachers. Classification 

of maternal smoking during pregnancy was obtained 

retrospectively at the time of child assessment from 

parental interview. Propensity score matching was used 

to create exposed and unexposed groups, which did 

not differ in their propensity to smoke with respect to 

16 characteristics. 

After matching, children exposed to maternal smoking 

during pregnancy were significantly more likely to score 

in the problematic range on the Strengths and Difficulties 

Questionnaire total difficulties index. Maternal smoking during 

pregnancy was more strongly associated with externalizing than 

internalizing behaviors. 

Gaysina et al. 

(2013)

• Examined three studies:

– Christchurch Health 

and Development 

Study, a longitudinal 

cohort study of 

adopted and biological 

children; 1,088 children 

reared by genetically 

related mothers and 

36 children reared by 

genetically unrelated 

adoptive mothers

– Early Growth and 

Development Study, 

a longitudinal study 

of adoption at birth; 

310 children reared by 

genetically unrelated 

adoptive mothers

– Cardiff IVF Study, an 

adoption-at-conception 

study among genetically 

related families and 

genetically unrelated 

families; 636 children 

reared by genetically 

related mothers and 

206 by genetically 

unrelated mothers

Outcomes included child conduct problems at 

4–10 years of age using the Rutter and Conners 

behavioral scales, the Child Behavior Checklist, the 

Children’s Behavior Questionnaire, and the Strengths 

and Difficulties Questionnaire. Regression analysis was 

used to test for dose–response relationships between 

maternal smoking during pregnancy and conduct 

problems in children. The potential confounders 

examined included the child’s gender, ethnicity, birth 

weight, and breastfeeding status; maternal age at birth 

of child and education; family socioeconomic status; 

family breakdown; placement age; and parenting 

practices. 

Among children reared by genetically related mothers and 

genetically unrelated mothers adopted at birth, a significant 

association was observed between maternal smoking during 

pregnancy and conduct problems in children. Children whose 

mothers smoked at least 10 cigarettes per day during pregnancy 

had the highest mean scores of conduct problems. In fully 

adjusted models, associations between maternal smoking during 

pregnancy and conduct problems in children were attenuated 

but remained significant for genetically related mother–child 

pairs and in genetically unrelated mother–child pairs in the 

Early Growth and Development Study, but they were not 

significant in the Christchurch Health and Development Study. 

Meta-analysis confirmed an association between maternal 

smoking during pregnancy and offspring conduct problems in 

genetically related mother–child pairs. 

Table A3.1-4 H Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-61

Study Design/population Outcomes examined Findings

O’Brien 

(2013)

• Pregnancy cohort study

• 176 offspring, 11–18 years 

of age, from the Maternal 

Infant Smoking Study of 

East Boston

The study examined potential interactions between 

prenatal exposure to maternal tobacco use and two 

functional genetic markers in the dopamine system to 

predict externalizing behaviors in offspring. Transporter 

gene DAT1 and dopamine receptor gene DRD4 were 

used to predict a latent composite of externalizing 

behavior by gender. Maternal tobacco exposure was 

assessed via questionnaire and serum cotinine levels 

during pregnancy; trimester-specific exposure was not 

included in the analysis. Behavioral outcomes included 

ADHD, oppositional defiance disorder, conduct disorder, 

substance use (nicotine, alcohol, and marijuana); and 

scales from the C-DISC-IV and the Antisocial Behavior 

Checklist. Multigroup regression analysis was conducted; 

covariates included maternal age, youth age, prenatal 

maternal alcohol and drug use, maternal and paternal 

antisocial behavior and harsh parenting, maternal 

employment, education, and total household income. 

The interaction between DAT1 (but not DRD4) and prenatal 

exposure to maternal tobacco use during pregnancy was 

significant in boys. DAT1 genetic susceptibility did not increase 

risk of externalizing behaviors in the absence of exposure to 

tobacco. The final model predicted 23% of the variance for girls 

and 60% of the variance for boys for externalizing behavior. Risk 

genotypes (DAT1) can modify the effects of prenatal exposure to 

tobacco in boys.

Jasinska et al. 

(2014)

• Review of existing 

literature of human 

neuroimaging research on 

nicotine and tobacco. 

This study examined current literature on several 

topics including: receptor-level PET and SPECT 

studies specific to nAChRs and upregulation of 

nAChRs inducted by chronic smoking; interaction of 

nicotine with the mesocorticolimbic dopamine system; 

functional activity and connectivity; and fMRI studies 

detailing the behavioral/cognitive effects of nicotine 

and large-scale brain networks.

This review summarized the literature that discusses the 

addictive properties and cognition-enhancing effects of nicotine; 

highlighted the clinical significance of the elucidation of 

neurobiological mechanisms of nicotine; and presented an 

overview of the use of PET, SPECT, and fMRI neuroimaging 

methods used to study nicotine in vivo, including the effects of 

nicotine on nAChRs and the dopamine system.

Kuja-Halkola 

et al. (2014)

• 2,754,626 children born 

in Sweden 

Data were obtained from several nationwide Swedish 

registries. Traditional and within-family analyses 

examined associations between maternal smoking 

during pregnancy and outcomes in offspring, including 

low academic achievement, general cognitive ability, 

and externalizing behaviors. Data on maternal 

smoking during pregnancy was collected during the 

first prenatal care visit. Outcomes for exposed and 

unexposed offspring were compared using linear and 

logistic regression; potential confounders included 

maternal age, birth year, and gender of offspring.

Through traditional analysis, maternal smoking during 

pregnancy was significantly associated with all outcomes. 

However, none of the associations remained significant after 

within-family analysis of discordant-exposed offspring. In 

quantitative genetic models, genetic factors explained the 

majority of the associations. 

Table A3.1-4 H Continued
A Report of the Surgeon General

A3.1-62 Appendix 3.1

Notes: ADHD = attention deficit hyperactivity disorder; CI = confidence interval; C-DISC-IV = Diagnostic Interview for Children−Version 4; CSI = Child Symptom 

Inventory; DISC-YC = Diagnostic Interview Schedule for Children-Parent Scale-Young Child; DSM-IV = Diagnostic and Statistical Manual of Mental Disorders, 4th Edition; 

fMRI = functional magnetic resonance imaging; ICD10 = International Statistical Classification of Diseases and Related Health Problems, 10th revision; LTP = long-term 

synaptic protentiation; nAChR = nicotinic acetylcholine receptor; NLSY = National Longitudinal Survey of Youth; NYS = New York State Follow-Up; OR = odds ratio; 

PET = positron emission tomography; SCAN = Screening test for Auditory Processing Disorders; SNP = single-nucleotide polymorphism; SPECT = single photon emission 

computed tomography; TEOAEs = transient evoked otoacoustic emissions.

Table A3.1-4 H Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-63

Table A3.1-5 Preclinical/animal studies on fetal nicotine exposure

Study Species/exposure Measures Findings Comments

Navarro et al. 

(1988)

• Pregnant Sprague-Dawley 

rats

• Osmotic minipumps 

delivered saline or nicotine 

(6 mg/kg/day) from 

gestational days 4 to 20 or 

twice-daily injections of saline 

or nicotine (3 mg/kg/day, s.c.) 

from gestational days 4 to 20.

• Catecholaminergic system 

function

• Tissue was collected at various time 

points postnatally (P2, P5, P10, P15, 

P22, P30, P35, and P45) for assessment 

of catecholamine levels and utilization 

rates.

• Minipump delivery of nicotine 

produced an overall reduction in NE 

and DA levels and use in the cerebral 

cortex immediately after birth that 

dissipated between postnatal weeks 

2 and 4 but became apparent again in 

young adulthood for NE levels only.

• There were no changes in cerebral NE 

synaptosomal uptake, but TH activity 

was decreased; in the midbrain and 

brainstem, lower NE levels and use 

were apparent in young adulthood 

only but without changes in TH or 

synaptosomal uptake. 

• Reduced NE and DA content was 

also seen in the kidneys and lungs, 

but repeated s.c. injections had a 

stimulatory effect on neurotransmitter 

levels and utilization rates.

—

Sorenson et al. 

(1991)

• Sprague-Dawley rats 

• Rats received nicotine (0 or 

~6 mg/kg/day) in drinking 

water starting 15 days before 

mating and throughout 

gestation.

• Attention, cognition—

radial eight-arm maze task

• Adolescent offspring (P45–P65) were 

tested in a radial eight-arm maze task. 

• Nicotine exposure impaired learning 

in both genders in the eight-arm 

maze.

Nicotine exposure during gestation 

impairs acquisition of an attentional 

learning task without changing levels of 

motivation, suggesting lower impulse 

control.
A Report of the Surgeon General

A3.1-64 Appendix 3.1

Study Species/exposure Measures Findings Comments

Richardson 

and Tizabi 

(1994)

• Pregnant Sprague-Dawley 

rats

• Osmotic minipumps 

delivered saline or nicotine 

(6 mg/kg/day) from 

gestational days 4 to 20.

• Catecholaminergic system 

function

• Early-adolescent offspring (P19–P21) 

were tested for hyperactivity and for 

DA and D2 receptor levels. 

• Gestational nicotine produced dosedependent hyperactivity, as measured 

by open-field locomotion in a subset of 

offspring.

• In these hyperactive offspring, DA 

levels were decreased in the VTA and 

striatum, but DA levels increased in 

the substantia nigra.

• DA and D2 receptor levels also were 

reduced in the striatum. 

• There were no changes in DA or D2

receptor levels in the PFC or NAc. 

In adult animals, nicotine-induced 

locomotion is attributable to activation of 

the mesolimbic DA pathway. These data, 

however, suggest that the nigrostriatal 

DA pathway contributes to hyperactivity 

in offspring of nicotine-exposed dams via 

increased striatal DA release. 

Slotkin et al. 

(1995)

• Pregnant Sprague-Dawley 

rats

• Osmotic minipumps 

delivered saline or nicotine 

(0.7 or 2.1 mg/kg/day free 

base) from gestational 

days 5 to 22.

• Response to hypoxia • 1-day-old pups were placed in hypoxic 

conditions (5% O2 balanced with N2).

• Survival, adrenal catecholamine 

release, and release of norepinephrine 

in the brain were assessed.

• 15% of the animals exposed to the 

high dose of prenatal nicotine died.

• Basal adrenal catecholamine release 

was unaffected by nicotine treatment, 

but normal norepinephrine release 

in response to hypoxia was absent in 

nicotine-exposed animals without 

changes in noradrenergic receptor 

binding.

Prenatal nicotine exposure leads to 

the loss of neonatal hypoxia tolerance. 

Two mechanisms likely contribute: 

cardiovascular, mediated by adrenal 

catecholamines; and respiratory, 

mediated by release of norepinephrine 

in the central nervous system. Prenatal 

nicotine exposure led to the complete 

loss of the adrenomedullary release in 

response to hypoxia. 

Table A3.1-5 Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-65

Study Species/exposure Measures Findings Comments

Fewell and 

Smith (1998)

• Pregnant Sprague-Dawley 

rats

• Osmotic minipumps 

delivered saline or nicotine 

(6 mg/kg/day free base) from 

gestational days 6 to P5 or P6.

• Response to hypoxia • On P5 or P6, pups were exposed 

to a single period of hypoxia 

(97% N2−3% CO2), and time to last 

gasp was recorded; or they were 

exposed to repeated hypoxic events and 

their ability to autoresuscitate from 

primary apnea was measured.

• Prenatal nicotine exposure had no 

effect on time to last gasp, but it 

significantly impaired the ability for 

autoresuscitation from primary apnea 

(18 ± 1 recovered periods in vehicleexposed pups vs. 12 ± 2 recovered 

periods in nicotine-exposed pups). 

• Autoresuscitation failure was 

attributable to A-V dissociation 

following early cardiac resuscitation.

Nicotine exposure during gestation 

impairs newborns’ ability to 

autoresuscitate from primary apnea 

during repeated hypoxic events, like what 

may happen during repeated episodes 

of sleep apnea. The mechanism may be 

altered non-neurogenic catecholamine 

release.

Table A3.1-5 Continued
A Report of the Surgeon General

A3.1-66 Appendix 3.1

Study Species/exposure Measures Findings Comments

Slotkin (1998) — • Review on disruption 

of neuron replication, 

differentiation, and 

function following 

gestational nicotine.

— Review article highlighting animal 

models of the effects of gestational 

nicotine on fetal neuron development 

and function. Emphasized findings 

are that prenatal nicotine increases 

biomarkers of cell damage and decreases 

cell number in fetal brain, likely through 

a persistent increase in constitutive 

cFos expression that activates apoptosis. 

Nicotine also decreases DNA synthesis 

and negatively affects adrenomedullary 

function. nAChR upregulation and 

resulting hypoactive cholinergic systems 

after nicotine exposure also likely 

contribute to behavioral abnormalities 

in exposed offspring. The relationship 

between these changes and negative 

consequences in offspring like SIDS 

or behavioral deficits is discussed. The 

greater impact of tobacco/nicotine on 

fetal development compared to cocaine is 

also discussed. 

Muneoka et al. 

(1999)

• Pregnant Sprague-Dawley 

rats

• Daily injections of saline or 

nicotine (3 or 6 mg/kg, s.c.) 

from gestational days 7 to 20 

• Tissue collected from male 

pups at 8–9 weeks

• Catecholaminergic system 

function 

• HPLC-ECD was used to measure DA, 

NE, serotonin, and their metabolites 

in the cortex, hippocampus, striatum, 

hypothalamus, and cerebellum.

• Exposed offspring trended toward 

lower DA in the cortex, but they had 

significantly lower DOPAC content in 

the cortex only. 

• No differences seen in NE, serotonin, 

HVA, or 5-HIAA.

High-dose nicotine during gestation 

reduces total cortical dopaminergic 

activity via reduced innervation. Authors 

suggest that this DA dysfunction may 

contribute to offsprings’ vulnerability to 

drug abuse and psychiatric disorders.

Table A3.1-5 Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-67

Study Species/exposure Measures Findings Comments

Aramakis et al. 

(2000)

• Male and female SpragueDawley pups were injected 

twice daily with saline or 

nicotine (1 or 2 mg/kg, 

s.c.) from P1 to P7–8, P8 to 

P11–14, or P20 to P23–25, 

and slices were prepared the 

next morning.

• A fourth group received saline 

or nicotine (2 mg/kg, s.c.) 

from P8 to P16, and slices 

were prepared 7–10 days later.

• Development of excitatory 

neurotransmission in the 

auditory cortex during a 

critical period 

• Whole-cell recordings were used 

to measure glutamate EPSPs and 

intrinsic membrane properties of layer 

II–IV pyramidal neurons.

• Nicotine exposure during postnatal 

weeks 1 or 4 had no effect on synaptic 

function. Exposure during postnatal 

week 2 produced lasting changes 

in EPSPs, with longer durations, 

multiple peaks, and enhanced NMDAR 

components, without changing 

number or binding of a7 nAChRs.

• Prior exposure to nicotine also blunted 

normal nicotine-enhanced NMDAR 

EPSPs.

• Membrane properties related to spike 

generation, but not passive membrane 

characteristics, were also altered by 

nicotine during postnatal week 2.

Normal maturation of nAChR regulation 

of glutamatergic neurotransmission in the 

auditory cortex is disrupted by nicotine 

exposure during the second, but not first or 

fourth, postnatal week. This disruption is 

lasting and selective to NMDAR-mediated 

neurotransmission and is not associated 

with changes in a7 nAChR number or 

binding. The second postnatal week may 

represent a critical period of nAChR 

development in the auditory cortex that 

can be disrupted by nicotine.

Klein et al. 

(2003)

• Pregnant C57Bl/6J mice

• Mice received 2% SAC alone 

or with nicotine (50 μg/mL) 

in drinking water from 

gestational day 9 to weaning 

of pups on P21.

• Response to drug rewards 

in later developmental 

stages—oral nicotine selfadministration

• Male and female offspring were tested 

during periadolescence (P35–42) for 

oral self-administration of nicotine.

• Animals had unlimited access to 

SAC-only and sweetened nicotine 

solution (50 μg/mL nicotine) in home 

cages in a two-bottle choice test, with 

bottles switched daily to prevent place 

preference.

• Exposed offspring consumed 

significantly more nicotine-containing 

solution than did unexposed offspring. 

• Further analyses found that exposed 

males had increased preference for 

nicotine over SAC (64.6% vs. 23.2%, 

respectively), while females did not 

(35% vs. 37.8%, respectively). 

Perinatal nicotine exposure produces 

gender-dependent increases in nicotine 

preference during the periadolescent 

period, with only males showing greater 

nicotine preference. This gender 

difference may be attributable to prenatal 

nicotine-induced nAChR upregulation 

occurring in males but not in females. 

Table A3.1-5 Continued
A Report of the Surgeon General

A3.1-68 Appendix 3.1

Study Species/exposure Measures Findings Comments

Pauly et al. 

(2004)

• C57Bl/6 mice

• Mice received 2% SAC alone 

or with nicotine (200 μg/mL 

free base) in drinking water 

starting 30 days before 

mating and until gestational 

day 18.

• Hyperactivity in later 

developmental stages—

open-field test

• Offspring exposed to prenatal SAC 

or nicotine were tested for basal 

locomotor activity at P20, P40, 

and P60, followed the next day by 

nicotine-induced (0 or 1.0 mg/kg, i.p.) 

hypothermia testing. 

• Female-exposed offspring displayed 

hyperactivity at P20 but no treatment 

differences at P40 or P60.

• Male-exposed offspring were 

hyperactive at P40 and P60 and 

displayed significant stereotypy 

compared with unexposed male 

offspring. 

• Females, but not males, showed 

a blunted hypothermic response to 

a nicotine challenge at P60.

Gestational nicotine gender-dependently 

influences hyperactivity and acute 

pharmacologic responses to nicotine 

during adolescence and early adulthood. 

Higher stereotypy in exposed male 

offspring may be attributable to nicotineinduced increases in dopaminergic 

activity.

Levin et al. 

(2006)

• Pregnant Sprague-Dawley 

rats

• Osmotic minipumps 

delivered water or nicotine 

(6 mg/kg/day free base) from 

gestational days 4 to 21.

• Response to drug rewards 

in later developmental 

stages—nicotine selfadministration with forced 

abstinence

• Female offspring underwent standard 

i.v. self-administration of nicotine 

(0.03 mg/kg/infusion free base) for 

4 weeks starting at P40–46. 

• A 7-day drug-free period followed the 

4 weeks of self-administration before 

self-administration was tested for 

5 days starting at P82.

• Initial acquisition of nicotine selfadministration was unaffected by 

prior nicotine exposure, but selfadministration was significantly 

increased in exposed animals after 

forced abstinence.

Prenatal nicotine produced a 

significant increase in i.v. nicotine selfadministration during adulthood that 

became apparent only after a period 

of forced abstinence from nicotine. 

It should be noted that offspring 

were single-housed after weaning. 

Isolation rearing can alter a multitude 

of behaviors, including enhanced 

psychostimulant self-administration 

(Hall et al. 1998).

Table A3.1-5 Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-69

Study Species/exposure Measures Findings Comments

Liang et al. 

(2006)

• Sprague-Dawley rats 

• Twice-daily injections of 

saline or nicotine (0.7 mg/kg 

free base, s.c.) from P8 to P12, 

when expression of nAChRs 

peaks in the primary auditory 

cortex

• Auditory processing—

nAChR regulation of 

signaling processing 

in primary cortex and 

auditory learning

• Adults (>P60) underwent auditorycued active avoidance and in vivo 

auditory physiology testing. 

• Perinatal nicotine significantly 

decreased avoidance behavior without 

affecting tone orienting or motor 

function.

• Acute nicotine exposure 

(0.7 mg/kg, s.c.) in control rats 

enhanced sensitivity to sound of 

neural responses in layers III and 

IV of A1 that was attenuated by 

the nonspecific nAChR antagonist 

mecamylamine.

• Exposed animals did not show this 

enhancement of auditory responses 

after acute nicotine.

nAChRs in the primary auditory cortex 

normally function to enhance sensitivity 

and responsiveness to auditory stimuli, 

but exposure to nicotine during postnatal 

week 2 impairs auditory learning. These 

deficits are not associated with changes 

in nAChR binding.

Pentel et al. 

(2006)

• Pregnant Holtzman SpragueDawley rats (with or without 

prior nicotine-vaccine 

exposure) received computercontrolled saline or nicotine 

(2 mg/kg/day free base, i.v.) 

from gestational days 1 to 20.

• On gestational day 20, a 

final dose of radiolabeled 

nicotine (0.05 mg/kg, i.v.) 

was delivered, and fetal 

blood/tissue was collected 

25 minutes later.

• Activation of placental 

nAChRs by maternal 

nicotine

• Gestational nicotine exposure 

produced significant increases in 

brainwide nAChR binding in the fetal 

brain and spinal cord, as measured by 

125I-epibatadine binding. 

• Nicotine exposure also decreased 

cFos induction in the fetal caudate, 

adrenal glands, and lungs, but not the 

hippocampus.

• The nicotine vaccine did not affect 

fetal receptor binding or cFos

expression.

Using a dosing regimen that models 

human smoking, gestational nicotine 

upregulates nAChR density throughout 

the fetal brain and spinal cord. Maternal 

vaccination against nicotine reduces 

nicotine distribution in the fetal brain, 

but it had no influence on nAChR density 

or neuronal activity. 

Table A3.1-5 Continued
A Report of the Surgeon General

A3.1-70 Appendix 3.1

Study Species/exposure Measures Findings Comments

Dwyer et al. 

(2008)

— • Review on the role of 

nAChRs in regulating early 

brain plasticity 

— Review article highlighting the profound 

and diverse contribution of nAChRs in 

regulating the proper maturation of 

neural circuitry, from influencing cell 

survival and patterned spontaneous 

activity to regulation of developmental 

switches in GABAergic signaling. 

Includes a commentary on animal 

models of gestational nicotine exposure 

and summarizes clinical and preclinical 

work on the effects of gestational 

nicotine exposure, with a focus on 

SIDS, behavioral disorders, and auditory 

processing deficits. 

Franke et al. 

(2008)

• Pregnant Sprague-Dawley 

rats

• Osmotic minipumps 

delivered saline or nicotine 

(3 mg/kg/day free base) from 

gestational days 4 to 22.

• Cocaine exposure for 

cFos analysis was done on 

postnatal day 32.

• Response to natural or 

drug rewards in later 

developmental stages—

sucrose or cocaine selfadministration

• Neural activation in 

response to cocaine

• Adolescent rats (P32–43) exposed to 

prenatal nicotine showed an overall 

decrease in motivation to consume 

sucrose compared with nonexposed 

animals.

• Both exposed and nonexposed animals 

acquired cocaine self-administration, 

but nonexposed animals worked more 

for the lower 200 μg/kg/infusion dose, 

while exposed animals worked more 

for the high 500 μg/kg/infusion dose.

Prenatal nicotine exposure has complex 

effects on responding to natural and drug 

rewards in adolescent offspring, with 

lower motivation for natural rewards 

and greater motivation for higher doses 

of cocaine, as compared with unexposed 

offspring. This may reflect decreased 

aversion in exposed offspring and is 

corroborated by less stereotypy to high 

doses.

Table A3.1-5 Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-71

Study Species/exposure Measures Findings Comments

Chistyakov et 

al. (2010)

• Pregnant outbred Swiss mice

• Mice received SAC alone or 

with nicotine (200 μg/mL free 

base) from 2 weeks before 

mating up to weaning of pups 

at P21.

• Response to drug rewards 

in later developmental 

stages—nicotine selfadministration

• Adolescent male mice (6 weeks 

old) self-administered saline or 

nicotine (0.028, 0.056, 0.084, or 

0.112 μg/infusion, free base) for one 

30-minute test.

• Exposed mice had significantly 

higher nicotine self-administration 

of 0.028 and 0.056 μg/infusion and 

similar levels of self-administration 

of the 0.084 μg/infusion dose than 

unexposed mice.

• Spontaneous locomotor activity was 

tested at weeks 3, 4, 5, 7, and 10, but 

nicotine exposure had no effect in 

novel or familiar environments.

• [3H]epibatidine-binding sites were 

unaffected by prenatal nicotine in 

the cortex and hippocampus at weeks 

3 and 6.

The reinforcing effects of nicotine are 

sensitized in adolescent offspring of 

nicotine-exposed mothers without 

influencing locomotor behavior 

or nAChR binding in the cortex or 

hippocampus. As a result, exposed 

offspring self-administer more nicotine at 

low doses than unexposed offspring do.

Table A3.1-5 Continued
A Report of the Surgeon General

A3.1-72 Appendix 3.1

Study Species/exposure Measures Findings Comments

Cao et al. 

(2011)

• Pregnant Sprague-Dawley 

rats

• Osmotic minipumps 

delivered saline or nicotine 

(3 mg/kg/day free base) from 

gestational days 4 to 18.

• Expression of genes 

involved in neuronal 

signaling and addiction

• Expression of 29 CAM genes associated 

with drug addiction was measured by 

qRT-PCR in exposed and unexposed 

female offspring (P35).

• Nicotine exposure modified various 

CAM genes of the neurexin, 

immunoglobulin, cadherin, and 

adhesion-GPCR superfamilies, as 

well as their intracellular signaling 

pathways in the CPu, NAc, amygdala, 

and PFC.

• Most of the affected CAM genes 

were downregulated, but Postn was 

upregulated in the NAc and PFC. 

• The CPu was particularly sensitive to 

modification by prenatal nicotine, with 

17 genes showing altered expression.

• Rho small GTPase-related pathways 

were modified by nicotine exposure in 

all four brain regions, while MAPKrelated pathways were altered only in 

the PFC and amygdala.

• GPCR-related signaling pathways, 

Notch and Wnt/Frizzled pathways, and 

growth factor-signaling pathways were 

also influenced by nicotine exposure in 

the NAc, PFC, and amygdala.

Gestational nicotine influences the 

expression of CAMs in a region-specific 

manner. In striatal regions, CAMs 

related to glutamate synapse structure 

and function were downregulated, 

with the CPu showing the greatest 

vulnerability. CAMs related to GABAergic 

synapse formation were altered in the 

PFC. Myelination-related CAMs were 

downregulated in both the PFC and CPu, 

suggesting that neuron–glia interactions 

are also affected by gestational nicotine. 

In the amygdala, CAMs important for 

emotional learning and memory were 

affected by nicotine exposure. Overall, 

prenatal nicotine likely reduces excitatory 

synapses in the ventral striatum while 

also compromising glutamatergic 

signaling in the dorsal striatum.

Table A3.1-5 Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-73

Study Species/exposure Measures Findings Comments

Wei et al. 

(2011)

• Pregnant Sprague-Dawley 

rats

• Osmotic minipumps 

delivered saline or nicotine 

(3 mg/kg/day free base) from 

gestational days 4 to 22.

• Expression of genes 

involved in neuronal 

signaling and addiction

• Female offspring (P35) of exposed or 

unexposed dams were used to assess 

cell survival/death, development/

plasticity, immune response, and 

cellular metabolism-related pathways 

in the PFC, caudate putamen, NAc, 

PVN, and the amygdala.

• The most robust changes occurred 

in cell death/survival pathways, with 

24 biological pathways related to 

cell death/survival modulated by 

gestational nicotine in a regiondependent manner. 

• These pathways were further classified 

into three categories: growth factor, 

death receptor, and kinase cascade.

• In the NAc, growth factor, kinase, and 

transcription factor expression were 

upregulated by prenatal nicotine, 

while growth factors, death receptors, 

and caspases were upregulated in the 

striatum by nicotine exposure. 

• Death receptor and caspase expression 

were decreased in the PFC.

Nicotine modulates both growth factor 

and death-receptor pathways by affecting 

the expression of their genes. The 

NAc showed an enhancement of cell 

survival pathways following nicotine 

exposure, while both cell death and 

survival pathways were increased in the 

structurally related striatum. Therefore, 

the balance of these pathways may 

determine the final effect of nicotine on 

cell survival.

Horst et al. 

(2012)

• Pregnant C57BL/6J mice 

• Mice were given nicotine 

(200 μg/mL) or vehicle 

(0.2% tartaric acid) in 

sweetened (2% SAC) drinking 

water throughout pregnancy 

and until pups were weaned 

at P21.

• Exposed offspring were 

behavior tested in adulthood 

and compared to drug-naïve 

β2 nAChR KO animals.

• Auditory processing—

auditory discrimination 

task

• Perinatal nicotine or lack of β2 

subunit disrupted learning of auditory 

discrimination task attributable to 

increased probability of false-alarm 

responses.

Perinatal nicotine impairs auditory 

processing in an appetitive task. 

Impairments are long-lasting, as testing 

was done months after end of drug 

exposure. Some potential mechanisms: 

nicotine during critical developmental 

periods: (1) interferes with β2 nAChR 

subunit expression or function that 

is necessary for normal auditory 

learning, and/or (2) disrupts function of 

thalamocortical neurons.

Table A3.1-5 Continued
A Report of the Surgeon General

A3.1-74 Appendix 3.1

Study Species/exposure Measures Findings Comments

Lacy et al. 

(2012)

• Pregnant Sprague-Dawley 

rats

• Rats received intermittent 

injections of saline or nicotine 

(0.05 mg/kg/injection) three 

times a day from gestational 

days 8 to 21.

• Response to natural 

rewards in later 

developmental stages—

sucrose self-administration

• Adult offspring of both genders were 

tested for habituation of spontaneous 

locomotor activity and for operant 

responding to a natural reward.

• Prior nicotine exposure had no 

effect on habituation to spontaneous 

locomotor activity.

• For operant experiments, offspring 

acquired FR3 responding for 

26% sucrose (w/v) before testing on 

varying concentrations (0%, 3%, 10%, 

30%, and 56%, Latin-square design) 

on an FR3 and PR schedule.

• Nicotine had no effect on acquisition 

of sucrose responding.

• Both exposed and nonexposed 

offspring showed an inverted U-shaped 

response curve, with peak responding 

for 10% sucrose.

• PR responding was significantly 

higher in nicotine-exposed rats than in 

nonexposed rats.

Nicotine exposure during gestation 

increases motivation for sucrose 

without altering sensitivity to varying 

concentrations of sucrose, weight, or 

spontaneous locomotor activity. These 

data suggest that prenatal exposure 

to low doses of nicotine results in 

long-term increases in motivation for 

natural rewards, which may help explain 

correlations between maternal smoking 

and adolescent obesity.

Schneider et 

al. (2012)

• Pregnant Wistar rats

• Rats received nicotine 

(0.06 mg/mL) in drinking 

water 3 weeks before mating 

and throughout gestation.

• Behavioral testing in male/

female offspring P25–P50

• Attention, five-choice 

serial reaction time task 

(5-CSRTT), locomotor 

behavior, and delaydiscounting

• Exposed offspring showed 

hyperactivity in a familiar 

environment; behavioral disinhibition 

in 5-CSRTT (i.e., more anticipatory 

responses, fewer omission errors, 

and shorter latency to errors); but no 

change in choice impulsivity.

Prenatal nicotine results in hyperactivity 

and greater impulsivity, which resemble 

some symptoms of ADHD. Previous work 

showed increased impulsivity in adult 

offspring with prenatal nicotine exposure, 

but symptoms of most psychiatric 

disorders emerge during adolescence.

Table A3.1-5 Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-75

Study Species/exposure Measures Findings Comments

Zhu et al. 

(2012)

• Pregnant C57BL/6 mice 

• Mice were given 2% SAC 

or nicotine (0, 0.05, 0.1, or 

0.2 mg/mL) plus 2% SAC in 

drinking water 3 weeks before 

mating and throughout 

pregnancy.

• High offspring mortality in 

the 0.2 mg/kg nicotine group 

precluded testing, and so this 

dose was not used further.

• Catecholaminergic system 

function

• P42 and P60 offspring were tested for 

hyperactivity, regional brain volume, 

radial thickness, and DA turnover.

• Offspring exposed to 0.1 mg/kg 

nicotine had significantly higher 

locomotor behavior in an open field 

than offspring exposed to SAC only or 

to 0.05 mg/kg nicotine.

• Spontaneous hyperactivity in the 

dark period and beginning of the 

light period was also seen in nicotineexposed offspring.

• Oral, but not intraperitoneal, 

methylphenidate (0.75 mg/kg) 

transiently decreased hyperactivity 

in nicotine-exposed offspring in 

comparison to control levels.

• Nicotine exposure significantly increased 

DA content but decreased (~100%) DA 

turnover (i.e., DOPAC/DA) in the PFC, 

and it had no effect on serotonin content 

or turnover.

• Striatal DA turnover was also 

decreased (~50%) by prenatal nicotine.

• Turnover was normalized by 

methylphenidate (0.75 mg/kg, p.o.).

• Decreased cingulate cortex volume 

and radial thickness were also a 

consequence of prenatal nicotine.

Gestational nicotine produces behavioral, 

neuroanatomical, and neurochemical 

changes in mice that parallel the human 

ADHD phenotype. These changes are 

limited to the dopaminergic system, 

and hyperactivity and DA turnover are 

normalized by oral methylphenidate 

treatment.

Table A3.1-5 Continued
A Report of the Surgeon General

A3.1-76 Appendix 3.1

Study Species/exposure Measures Findings Comments

Cao et al. 

(2013)

• Pregnant Sprague-Dawley rats

• Osmotic minipumps delivered 

nicotine (3 mg/kg/day free 

base) from gestational 

days 4 to 18.

• PFC, NAc, and CPu were 

collected from pups at 

P20–21, P35–36, or P59–60.

• White matter structure • Exposed adolescent (P35–36) males 

had increased myelination in PFC, 

CPu, and NAc, while females had 

decreased myelination in PFC, no 

change in CPu, and some genes 

upregulated in NAc.

• Levels normalized by adulthood for 

males but not for females.

Prenatal nicotine age-, gender-, and 

region-dependently alters myelination. 

Juvenile and adolescent gender 

differences may be attributable to 

gonadal hormone stimulation of 

oligodendrocyte development and 

subsequent myelination. Lasting changes 

in females may suggest greater sensitivity 

to the effects of gestational nicotine.

Chang et al. 

(2013)

• Pregnant Sprague-Dawley 

rats

• Osmotic minipumps 

delivered nicotine (0.075 or 

1.5 mg/kg/day) for 14 days.

• Nicotine delivery started on 

gestational day 8. 

• Consummatory behavior—

oral intake of alcohol, 

nicotine, and high-fat diet

• Oral consumption of ethanol, nicotine, 

and a high-fat diet were tested in 

adolescent offspring (P40–60).

• Prenatal nicotine exposure increased 

consumption of 0.02% nicotine 

solution, 2% ethanol solution, and a 

50% fat content chow without altering 

normal food and water intake.

Low-dose nicotine exposure during 

gestation increases consummatory 

behavior during adolescence that may 

lead to obesity. The mechanism is 

likely induction of orexigenic peptide 

systems in the hypothalamus and central 

amygdala.

Gorini et al. 

(2013)

• Pregnant Sprague-Dawley 

rats

• Osmotic minipumps 

delivered saline or nicotine 

(6 mg/kg/day free base) 

for 28 days, starting from 

gestational day 3.

• Brainstem function • Tissue from pups (ages P3–7) of both 

genders was used for assessment of 

TCR.

• Nicotine-exposed pups had 

significantly greater TCR than 

unexposed pups (average decrease 

in heart rate of 71 ± 12.4 bpm vs. 

35 ± 2.6 bpm).

• Nicotine exposure also increased 

trigeminally evoked glutamate 

neurotransmission by 64.2% ± 7.2% 

compared with unexposed pups.

• Serotonin 5-HT1A and 5-HT2A/C

antagonists attenuated increases in 

EPSCs.

Prenatal nicotine enhances TCR and 

facilitates glutamatergic input to cardiac 

vagal nerves upon trigeminal afferent 

stimulation. This enhancement is 

attributable to increases in serotonergic 

signaling in the brainstem via 5-HT1A and 

5-HT2A/C receptors. Exaggerated TCR and 

greater glutamatergic neurotransmission 

to cardiac vagal neurons likely produce 

increased bradycardia, drops in blood 

pressure, and apnea that lead to SIDS. 

Table A3.1-5 Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-77

Study Species/exposure Measures Findings Comments

Morgan et al. 

(2013)

• Pregnant Sprague-Dawley 

rats

• Rats received thrice-daily 

injections of saline or 

nicotine (0.15 mg/kg/day, i.v.) 

from gestational days 8 to 20.

• Orexin/hypocretin 

expression and function

• Expression of orexin and MCH 

neurons in the LH and orexin 

projections from LH to VTA was 

measured in adult offspring (P130) of 

exposed and unexposed dams.

• Prenatal nicotine exposure increased 

the number of orexin neurons in the 

LH without affecting the expression of 

MCH neurons. 

• Almost all VTA dopaminergic neurons 

received contacts from orexin fibers.

• Prenatal nicotine significantly 

increased the number of orexinpositive appositions onto DA cells. 

Gestational nicotine produces longlasting increases in the number of LH 

orexin neurons and orexin-dopaminergic 

connections in the VTA. These data 

suggest that a greater orexin-mediated 

influence on motivation may mediate 

some of the behavioral effects seen 

in nicotine-exposed offspring. The 

mechanism may be that prenatal nicotine 

interferes with normal postnatal pruning, 

resulting in more LH orexin neurons, 

possibly through drug-induced increases 

in brain-derived neurotrophic factor.

Mychasiuk et 

al. (2013)

• Pregnant Long-Evans rats

• Daily injections of saline or 

nicotine (0.3 mg/kg, s.c.) after 

mating and until pups were 

born

• Neuronal morphology—

Golgi-Cox analysis of 

dendritic length, dendritic 

branching, and spine 

density 

• Golgi-Cox analysis was done in adult 

offspring (P100). 

• Exposed males had greater dendritic 

complexity, while females had less 

dendritic complexity in PFC and NAc. 

• In the orbital frontal cortex, all 

offspring had greater spine density, 

and males had more dendritic 

branching.

Prenatal nicotine induces long-lasting 

changes in neuronal morphology in 

regions critical for executive function 

and reward behavior. Authors suggest 

this may be a mechanism for gestational 

nicotine exposure’s increasing 

vulnerability to nicotine addiction later 

in life. 

Bailey et al. 

(2014)

• Pregnant WT or α5 nAChR 

subunit KO mice

• Mice received nicotine 

(200 μg/mL) in sweetened 

(+2% SAC) water throughout 

gestation and until weaning 

at P21.

• Tissue collected at postnatal 

week 3 or in adulthood.

• Neurochemical changes 

(acetylcholine)

• Whole-cell recording and 

immunohistochemistry of 

layer VI mPFC neurons 

• Exposed young and adult WT mice had 

greater apical dendritic complexity and 

decreased nAChR signaling.

• This pattern was also seen in 

unexposed α5 KO mice.

• Exposed KO mice showed normalized 

morphology but were more sensitive 

to acute nicotine, showing greater 

reduction in nAChR signaling.

Prenatal nicotine leads to persistent 

changes in apical dendrite morphology 

and physiology that are dependent, in 

part, on the α5 nAChR subunit. This 

implies that prenatal nicotine can 

alter dendritic morphology in layer VI 

mPFC neurons important for attention, 

although behavioral effects were not 

tested here.

Table A3.1-5 Continued
A Report of the Surgeon General

A3.1-78 Appendix 3.1

Study Species/exposure Measures Findings Comments

Lacy et al. 

(2014)

• Pregnant Sprague-Dawley 

rats

• Rats received intermittent 

injections of saline or nicotine 

(0.05 mg/kg/injection) three 

times a day from gestational 

days 8 to 21.

• Response to drug rewards 

in later developmental 

stages—methamphetamine 

self-administration

• Adult offspring (P70) were tested for 

methamphetamine self-administration 

on FR (0.05 mg/kg/infusion, i.v.) 

and PR (0.005, 0.025, and 

0.05 mg/kg/infusion, i.v.) schedules.

• Both genders acquired 

methamphetamine self-administration 

on an FR schedule regardless of 

prenatal nicotine exposure, but 

females acquired the behavior more 

rapidly.

• In PR responding, nicotine-exposed 

offspring of both genders had higher 

break points at all doses and had 

significantly higher intake at the 

0.025 and 0.05 mg/kg doses.

Gestational nicotine produces 

lasting increases in motivation for 

methamphetamine without influencing 

acquisition of methamphetamine selfadministration.

McNair and 

Kohlmeier 

(2015)

• Pregnant mice (from the 

Naval Medical Research 

Institute) 

• Mice received 2% SAC alone 

or with nicotine (300 μg/mL 

free base) in drinking water 

throughout gestation.

• Glutamatergic signaling in 

brain reward circuitry—

calcium imaging

• Calcium imaging was used to measure 

glutamatergic neurotransmission in 

the laterodorsal tegmental nucleus of 

offspring, P7–22. 

• Nicotine exposure led to decreased 

AMPA receptor-mediated calcium 

responses and a second-to-first 

response ratio >1 after a second 

application of AMPA. 

• In control animals, the second-to-first 

response ratio with AMPA stimulation 

was <1.

• Responses to repeated NMDA also 

showed a significant increase in 

nicotine-exposed offspring compared 

with unexposed offspring, without 

differences in calcium responses to a 

single NMDA application.

Gestational nicotine can reduce 

both AMPAR- and NMDAR-mediated 

glutamatergic neurotransmission in the 

laterodorsal tegmental nucleus, a brain 

region critical for assigning motivational 

salience. Decreased AMPA-mediated 

calcium responses are likely attributable 

to changes in receptor quantity, 

subunit composition, and/or calcium 

permeability.

Table A3.1-5 Continued
E-Cigarette Use Among Youth and Young Adults

Evidence Tables A3.1-79

Study Species/exposure Measures Findings Comments

Zhu et al. 

(2014)

• C57Bl/6 mice

• Mice received 2% SAC alone 

or with nicotine (0.1 mg/mL) 

in drinking water starting 

3 weeks before mating and 

throughout pregnancy.

• F1 females from PNE and 

SAC groups were bred with 

WT, drug-naïve males at 

6–8 weeks of age to generate 

the F2 generation.

• Females of the F2 generation 

were bred with WT, drugnaïve males at 6–8 weeks 

of age to produce the 

F3 generation. 

• A parallel set of experiments 

had F1 males from PNE and 

SAC groups mate with WT, 

drug-naïve females to produce 

the F2 generation.

• Hyperactivity in later 

developmental stages—

multigenerational effects 

of nicotine exposure 

and amelioration by 

methylphenidate 

• Spontaneous locomotor activity was 

assessed in 6- to 7-week-old F2 and F3 

male and female mice.

• F2 male and female mice derived 

from F1 PNE females and F3 male 

and female mice derived from F2 PNE 

females displayed significant increases 

in locomotor activity compared with 

SAC counterparts.

• Oral methylphenidate (0.75 mg/kg) 

decreased locomotor activity in F2 and 

F3 PNE mice.

• Locomotor activity of F2 mice derived 

from male F1 PNE mice was not 

significantly different from their 

SAC counterparts.

Nicotine-induced hyperactivity can be 

transmitted through the maternal line to 

affect offspring of both genders, and oral 

methylphenidate at clinically relevant 

doses can attenuate hyperactivity in 

these mice. These data suggest that 

the hypodopaminergic state present in 

F1 females was likely transmitted to the 

F2 and F3 generations. The mechanism 

for this change is likely epigenetic and 

may be limited to epigenetic changes of 

mitochondrial genes. See Leslie (2013) 

for further discussion on the implications 

of these epigenetic changes.

Note: A1 = primary auditory cortex; ADHD = attention deficit hyperactivity disorder; AMPA = α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid; AMPAR = AMPA 

receptor; A-V = atrial-ventricular; bpm = beats per minute; CAM = cell-adhesion molecule; cFos = protooncogene and immediate early gene used as a marker of neuronal 

activity; 5-CSRTT = 5-choice serial reaction time task; CO2 = carbon dioxide; CPu = caudate putamen; DA = dopamine; D1R = dopamine D1 receptor; DOPAC= 3,4dihydroxyphenylacetic acid, metabolite of dopamine; EPSCs = excitatory postsynaptic currents; EPSPs = excitatory postsynaptic potentials; F1 = first filial generation 

(and similar); FR = fixed ratio; G20 = gestational day 20 (and similar); GABAergic = any cell, especially any neuron, that releases gamma-aminobutyric acid 

(GABA); GPCR = Gprotein-coupled-receptor; GTPases (singular GTPase) are a large family of hydrolase enzymes that can bind and hydrolyze guanosine triphosphate 

(GTP); 5-HIAA = 5-hydroxyindole acetic acid, the primary metabolite of serotonin; HPLC-ECD = high-performance liquid chromatography electrochemical detection; 

5HT1AR = serotonin (5-hydroxytryptamine) receptor 1A; 5-HT2A/C = serotonin (5-hydroxytryptamine) receptor 2 A/C; HVA = homovanillic acid, a metabolite of dopamine; 

i.p. =intraperitoneal; i.v. = intravenous; kg = kilogram; KO = knockout; LH = lateral hypothalamus; MAPK = mitogen-activated protein kinase; 

MCH = melaninconcentrating hormone; mg = milligram; min = minute; μg = microgram; mL = milliliter; mPFC = medial prefrontal cortex; N2 = nitrogen; 

NAc = nucleus accumbens; nAChRs = nicotinic acetylcholine receptors; NE = norepinephrine; NMDAR = N-methyl-d-aspartate receptor; O2 = oxygen; P3 (and similar) 

= postnatal day 3; PFC = prefrontal cortex; PNE = prenatal nicotine exposure; p.o. = per os (by mouth); Postn = periostin, osteoblast-specific factor; PR = progressive 

ratio; PVN = paraventricular nucleus of the thalamus; qRT-PCR = quantitative real-time polymerase chain reaction; SAC = saccharin; s.c. = subcutaneous; 

SIDS = sudden infant death syndrome; TCR = trigeminocardiac reflex; TH = tyrosine hydroxylase; VTA = ventral tegmental area; Wnt = Wingless-Type MMTV Integration 

Site Family, member 1, intracellular signaling factors; WT = wild type; w/v = weight/volume.

Table A3.1-5 Continued
A Report of the Surgeon General

A3.1-80 Appendix 3.1

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